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Recently, orexin (ORX)ergic system has gained great attention for its major neuroregulatory role on fish motor, circadian, feeding activities and above all during water toxic conditions. Fish are particularly sensitive to acute sublethal copper (Cu) concentrations while little is known about neurobehavioral data after chronic Cu exposure. For this work, the marine teleost ornate wrasse (Thalassoma pavo) was monitored for 21 days during exposure to a relatively low CuCl2 concentration (0.25 mg/l). In particular, at 1 day fish displayed moderate reductions (−32%) of feeding behaviors versus controls that decreased up to −94% at 21 days. Swimming activities were also moderately (−44%) reduced at 1 day, which diminished even further (~ −75%) from 7 to 21 days. The lethargic attitudes seemed to be frequently interrupted by aquatic surface respiration and abnormal motor behaviors such as rapid and abrupt changes of direction. Contextually, an upregulatory trend of HSP90 mRNAs occurred especially in the lateral part of the dorsal telencephalon (Dl; +72%), medial preglomerular nucleus (NPGm; +87%), and torus longitudinalis (TLo; +108%). Moreover, some of these nuclei displayed extensive argyrophilic signals that are typical of notable neurodegenerative events. Interestingly, a single intraperitoneal administration of ORX-A (10 ng/g of body weight) at 21 days attenuated not only some behavioral impairments but also HSP90 mRNA levels and neurodegenerative events. Behavioral indications of the present results tend to underlie the valuable role of HSP90 transcript together with the ORXergic system as probable rescuing factors operating in marine fish during persistent adverse environmental states with eventual bearings on human health issues.

Emerging studies are beginning to suggest that emotional states together with healthful measures constitute pertinent features of our lifestyle in which bad eating habits but more importantly what our gut has to host are causing great concern. It is well known that humans have established mutual relationships with a wide array of colonized microbes (collectively called gut microbiota) consisting of bacteria, fungi, eukaryotic parasites and viruses. The gut microbiota has exhibited a notable ability of communicating with the brain via a two-way system that includes the vagus nerve, immune sites, and a number of neurotransmitters. Interestingly, stressful along with obesity, cognitive, and brain developmental states are strongly influenced by microbiota homeostatic conditions. It was our aim to investigate behavioral and obesity effects evoked by treatment with probiotics via neuroinflammatory factors and namely IL-1β, NLRP3, Caspase-1 and NF-kB levels in the Syrian golden hamster. Following treatment with a high-fat diet (HFD), in the presence or absence of a multi-species probiotic formulation, hamsters were exposed to an unpredictable chronic mild stress (UCMS) test for 4 weeks. Independently of the diet, probiotics treatment markedly reduced stress-like behaviors in the different mazes. Moreover, probiotics decreased hypothalamic expression levels of the pro-neuroinflammatory factors like IL-1β, NLRP3, Caspase-1 and NF-kB, whereas HFD increased them. Contextually, they decreased plasmatic levels of IL-1β, NLRP3 and caspase-1 but not NF-kB. Our findings clearly support probiotics as a potentially valuable treatment strategy in obesity and anxiety, thereby proposing them for clinical treatments in patients with metabolic and mood disorders.

The mechanisms by which inflammation affects the different emotional moods are only partially known. Previous works have pointed to stress hormones like glucocorticoids plus the vascular factor endothelin-1 as key factors evoking stressful states especially in relation to endothelial dysfunctions. With this work, it was our intention to establish the role of pro- and anti-inflammatory cytokine expression variations towards depression-like behaviors and consequently the development of neurodegeneration events caused by endothelial damages in the hamster (Mesocricetus auratus). Such a rodent, which is considered a valuable animal model to test depression and anxiety states, exhibited a variety of depression-like behaviors including reduction in sucrose consumption, locomotion, and exploration (p

It is well established that the maintenance of energy expenditure is linked to active hypothalamic neural mechanisms controlling adaptive stimuli such as food intake. Variations of glucose levels and hormonal (leptin plus orexin-A) parameters, which are involved with energy homeostasis during different behavioral states, have not yet been fully defined. In this first study, behavioral analyses of an unpredictable stress model dealing with the actions of a sub-chronic administration of orexin-A (ORX-A) and the anti-hunger neuropeptide, i.e., leptin (LEP) within the hypothalamic suprachiasmatic (SCH) nucleus, were conducted on the valuable hibernating rodent (hamster; Mesocricetus auratus) model noted for its distinct depression and anxiety states. Treatment with LEP accounted for a notable reduction (p

The various physiological alterations caused by copper (Cu) exposure in fish have attracted great interests toward neuronal strategies against Cu toxicity. Here, neurobehavioral activities (including anxiety-like behaviors) and transcriptional levels of heat shock protein (Hsp)90 and hypoxia inducible factor-1alpha (HIF-1α) were evaluated in the crucian carp (Carassius carassius) treated with nominal sub-lethal higher (1.45mg/L) and lower (0.30mg/L) concentrations of CuCl2. Both concentrations accounted for diminished swimming speed and food intake plus a strong preference for the dark side of the light/dark apparatus together with a reduction of crossings between the two compartments with respect to controls. Contextually, Hsp90 and HIF-1α transcripts were largely down- and up-regulated, respectively, in some brain areas such as in the medial part of the dorsal telencephalon (Dm, -52%) and in the ventral part of the ventral telencephalon (Vv, +68%). When carps were transferred to CuCl2-free water, some behaviors were rescued especially for fish previously exposed to 0.30mg/L concentration. In this same condition, Hsp90 mRNA levels were recuperated (-94%) in the medial preglomerular nucleus (NPGm) with respect to exposed fish while HIF-1α mRNA was mostly down-regulated in telencephalic stations. Moreover, recovery capacities of this extraordinary resistant fish was exhibited by evident reductions (-80%) of the dark argyrophilic granules such as in the ventral telencephalon (VTel). Overall, our results point to interesting responses against Cu toxicity involving Hsp90 and HIF-1α transcripts that may constitute early indicators of environmental stressors leading to the repair of metal-induced damages in fish with notable brain plasticity properties.

Recent indications are suggesting that high fat and sugar-enriched foods do not only evoke harmful physiological conditions, but they also endure evident structural alterations in cerebral regions controlling cognitive and feeding behaviors. Food consumption plus neuronal energy regulatory mechanisms seem to constitute a complex system assuring that food calories do not exceed body requirements. At the same time obesogenic-related properties of limbic feeding stations like the hypothalamus (HTH), hippocampus (HIP) and amygdala (AMY) tend to control eating habits through the interaction of distinct neuropeptides. For this purpose, it was our intention to correlate expression differences of a key anti-obesogenic neuropeptide receptor i.e. neurotensin1 (NTR1) on mnemonic performances in the hibernating hamster (Mesocricetus auratus) exposed to a high fat diet (HFD). Interestingly, these hamsters exhibited a notable enhanced (p < 0.01) body weight from the fifth on to the twelfth week of treatment, which was accompanied by elevated blood lipid cholesterolo and triglycerides and glucose levels. At the same time these hamsters provided diminished locomotor activities such as exploratory bouts, rearing and grooming behaviors. Of greater relevance was their very extreme (p < 0.001) inability of identifying new objects during novel object recognition (NOR) tests along with not having correctly chosen the chamber of the conditioned place preference (CPP) apparatus, which contained the gratifying reward. Surprisingly the altered behavioral plus mnemonic tasks of HFD hamsters were tightly related to elevated NTR1 expression changes in the above limbic sites thus proposing this neuronal system as a highly probable alternative for treating obesity-dependent mnemonic dysfunctions.

Food intake ensures energy resources sufficient for basic metabolism, immune system and reproductive investment. It is already known that food-seeking performances, which are crucially controlled by orexins (ORXs), may be under the influence of environmental factors including pollutants. Among these, mancozeb (mz) is becoming an environmental risk for neurodegenerative diseases. Due to few studies on marine fish exposed to mz, it was our intention to correlate feeding latency, food intake and feeding duration to potential neurodegenerative processes in key diencephalic sites and expression changes of the ORX neuroreceptor (ORXR) in the ornate wrasses (Thalassoma pavo). Hence, fish exposed for 4 days (d) to mz 0.2 mg/l (deriving from a 0.07, 0.14, 0.2, 0.3 mg/l screening test) displayed a significant reduction (p 0.05) of food intake compared to controls as early as 1d that became more evident (p 0.01) after 3d. Moreover, significant enhancements of feeding latency were reported after 1d up to 3d (p 0.001) and even feeding duration was enhanced up to 3d (p 0.001), which instead moderately increased after 4d (p 0.05). A reduction (-120%; p 0.001) of mean body weight was also detected at the end of exposure. Likewise, a notable (p 0.001) activation of ORXR protein occurred together with mRNA up-regulations in diencephalic areas such as the diffuse nucleus of the inferior lobe (+48%) that also exhibited evident degenerative neuronal fields. Overall, these results highlight an ORX role as a vital component of the neuroprotective program under environmental conditions that interfere with feeding behaviors.

Interaction of the orexinergic (ORXergic) neuronal system with the excitatory (glutamate, l -Glu) or the inhibitory (GABA) neurosignaling complexes evokes major homeostatic physiological events. In this study, effects of the two ORXergic neuropeptides (ORX-A/B) on their receptor (ORX-2R) expression changes were correlated to feeding and grooming actions of the hibernating hamster ( Mesocricetus auratus ). Infusion of the central amygdala nucleus (CeA) with ORX-A caused hamsters to consume notable quantities of food, while ORX-B accounted for a moderate increase. Interestingly the latter neuropeptide was responsible for greater frequencies of grooming with respect to both controls and the hamsters treated with ORX-A. These distinct behavioral changes turned out to be even greater in the presence of l -Glu agonist (NMDA) while the α 1 GABA A receptor agonist (zolpidem, Zol) greatly reduced ORX-A-dependent feeding bouts. Moreover, ORX-A + NMDA mainly promoted greater ORX-2R expression levels with respect to ORX-A-treated hamsters while ORX-B + Zol was instead largely responsible for a down-regulatory trend. Overall, these features point to CeA ORX-2R sites as key sensory limbic elements capable of regulating eating and grooming responses, which may provide useful insights regarding the type of molecular mechanism(s) operating during feeding bouts.

Hypothermia is a physiological condition assuring brain protection against hypoxic-related damages. In this con- text, investigations carried out on the facultative hibernator Mesocricetusauratusprovedtobeveryusefulfor establishing thetypeofneurosignalingroleexertedbycerebral α-amino-3- hydroxy-5-methyl-4-isoxazole-propionic acid receptor (AMPAR) subtypes during hypoxic/reperfusion injuries of the entrance (EN), torpor (TORP), and arousal (AROU) hi- bernating states. From the evaluation of the major AMPARs (GluR1 and GluR2), together with their scaffold proteins synapse-associated protein 97 kDa (SAP97) and PICK1, it resulted that GluR1 and SAP97 were mostly upregulated during the hypotensive (EN and TORP) states of the brainstem, amygdala, and hypothalamus, sites which are im- plicated with cardiovascular, motor, and sleep-wake events. In addition, elevated expression densities of the pro-apoptotic factor Bcl-2-associated X protein (Bax) resulted to be corre- lated to marked amino-cupric-silver stain signals during both hibernating states. Conversely, an increase of the neuroprotec- tive factor GluR2, together with PICK1 and the anti-apoptotic B cell lymphoma 2 (Bcl-2), appeared to be linked with re- duced argyrophilic signals in most of the above areas of the hypertensive AROU state. These first indications highlight distinct protective/degenerative measures of the above factors constituting key on/off switches during the various hibernat- ing states that may provide potential therapeutic bearings on sleeping disorders.

Several studies have pointed to the amygdala as a main limbic station capable of regulating different stressful states such as anxiety and depression. In this work it was our intention to determine the role of the central amygdala nucleus (CeA) on the execution of either anxiolytic and/or anti-depressant behaviors in the hibernating hamster (Mesocricetus auratus) via infusion of CeA with the antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) specific for α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) plus the specific agonist for α4 GABAAR i.e. 4,5,6,7-tetrahydroisoxazolo(5,4-c)pyridin-3-ol (THIP). Treatment with CNQX appeared to mainly prompt anti-depressant effects as shown by the achievements of swimming feats during forced swim test while THIP prevalently accounted for evident bouts of climbing when exposed to the same test. Moreover, even in the presence of the concomitant administration of both of these compounds, hamsters continued to spend more time in swimming despite this significant behavioral effect resulted to be numerically reduced for hamsters treated with only the α4 GABAAR agonist. Conversely, when these animals were tested in elevated plus maze (EPM), THIP tended to mostly favor anxiolytic activities as exhibited by stressed animals spending more time entering and remaining in EPM open arms. It was interesting to note that behavioral changes induced by both drugs appeared to be also responsible for glutamate receptor (GluR) expression differences as indicated by CNQX favoring an evident up-regulation of GluR2-containing neurons whereas THIP induced an up-regulation, this time of GluR1-containing neurons. Overall, the anti-depressant role of CNQX seems to be mostly attributed to elevated GluR2 levels while an anxiolytic-like effect of THIP was correlated to high GluR1values thereby proposing distinct GluRs as useful therapeutic sites against degenerative diseases such as depression-like behaviors.

It's known that neurons in mammalian hibernators are more tolerant to hypoxia than those in non-hibernating species and as a consequence animals are capable of awakening from the arousal state without exhibiting cerebral damages. In addition, evidences have suggested that euthermic hamster neurons display protective adaptations against hypoxia, while those of rats are not capable, even though molecular mechanisms involved in similar neuroprotective strategies have not been yet fully studied. In the present work, overstimulation of glutamatergic receptors NMDA recognized as one of the major death-promoting element in hypoxia, accounted for altered network complexity consistent with a moderate reduction of hippocampal neuronal survival (p < 0.05) in hamsters. These alterations appeared to be featured concomitantly with altered glutamatergic signaling as indicated by significant down-regulation (p < 0.01) of NMDAergic (NR2A) and AMPAergic (GluR1, R2) receptor subtypes together with the metabotropic mGluR5 subtype. Diminished mRNA levels were also reported for NMDA receptor binding factors and namely PSD95 plus DREAM, which exert positive and negative regulatory properties, respectively, on receptor trafficking events. Conversely, involvement of glutamatergic signaling systems on neuronal excitotoxicity was strengthened by the co-activation of GABA(A)R-mediated effects as indicated by toxic morphological effects being notably reduced along with up-regulated GluR1, GluR2, mGluR5, DREAM, and Homer1c scaffold proteins when muscimol was added. Overall, these results point to a neuroprotective role of the GABAergic system against excitotoxicity episodes via DREAM-dependent inhibition of NMDA receptor and activation of AMPA receptor plus mGluR5, respectively, thus proposing them as novel therapeutic targets against cerebral ischemic damages in humans.

At date, studies are beginning to show that ionotropic glutamate receptors (iGluRs) are influenced by the orexin/hypocretin (ORX) system during the regulation of several physiological functions in mammals. In this study, we showed that the effects of ORX-A and specific glutamatergic antagonists (3-(+)-2-carboxypiperazin-4-yl-propyl-1-phosphonate, CPP/cyano-7-nitro-quinoxaline-2,3-dione, CNQX) on the expression of NMDAR (NR1) and AMPAR (GluR2) subtype genes underlie a tight relationship of the ORXergic system with iGluRs in the teleost Carassius auratus (goldfish; Linnaeus, 1758). The application of specific primers for NR1 and GluR2 permitted us to identify two partial coding sequences (cds) of 115 bp and 90 bp that were highly similar (97% and 81%, respectively) to the related sequences of the rat (Rattus norvegicus). These cds were used to carry out a semi-quantitative RT-PCR (reverse transcription-polymerase chain reaction) in which a differentiated expression pattern of NR1 and GluR2 was obta...

Depletion of oxygen and glucose even for brief periods is sufficient to cause cerebral ischemia, which is a predominant worldwide cause of motor deficits with the reduction of life quality and subsequently death. Hence, more insights regarding protective measures against ischemic events are becoming a major research goal. Among the many neuronal factors, N-methyl-D-aspartate receptors (NMDAR), orexinergic neuroreceptors (ORXR), and sympatho-inhibitory neuropeptide catestatin (CST) are widely involved with ischemic episodes. In this study, it was possible to induce in vitro ischemic conditions of the hamster (Mesocricetus auratus) hippocampal and hypothalamic neuronal cultures, grown on a newly compartmentalized membrane system, via oxygen and glucose deprivation (OGD). These cultures displayed notably differentiated NMDARergic and ORXergic receptor expression activities along with evident brain-derived neurotrophic factor (BDNF) plus orexin A (ORX-A) secretion, especially under co-cultured conditions. Interestingly, addition of CST in OGD-insulted hippocampal cells accounted for upregulated GluN1 and ORX1R transcripts that in the case of the latter neuroreceptor was very strongly (p

This study focuses on the development of an advanced in vitro biohybrid culture model system based on the use of hollow fibre membranes (HFMs) and hippocampal neurons in order to promote the formation of a high density neuronal network. Polyacrylonitrile (PAN) and modified polyetheretherketone (PEEK-WC) membranes were prepared in hollow fibre configuration. The morphological and metabolic behaviour of hippocampal neurons cultured on PAN HF membranes were compared with those cultured on PEEK-WC HF. The differences of cell behaviour between HFMs were evidenced by the morphometric analysis in terms of axon length and also by the investigation of metabolic activity in terms of neurotrophin secretion. These findings suggested that PAN HFMs induced the in vitro reconstruction of very highly functional and complex neuronal networks. Thus, these biomaterials could potentially be used for the in vitro realization of a functional hippocampal tissue analogue for the study of neurobiological functions and/or neurodegenerative diseases.

The excessive levels of aquatic endocrine disruptors (EDs) and namely heavy metals plus xenoestrogens account for irregular gas exchange processes, reduced reproductive success, as well as abnormal social interactions of marine teleost fish. These effects at the encephalic level appear to derive from the interference of major signaling factors such as histamine (HA) neuroreceptor subtypes (H(1-4)R). HA is one of the main biogenic amine neuronal system responsible for regulatory homeostatic functions, including sleep-wake rhythms and motor activities. Recently, interests have begun to focus attention on toxic effects of some heavy metals, i.e., cadmium (Cd) and lead (Pb), and how they are capable of eliciting motor dysfunctions via HAergic receptor subtypes. Interestingly, subtype 2 (H(2)R) proved to be a preferential target of heavy metal-dependent altered locomotor maneuvers, as displayed by its specific antagonist (cimetidine)-inducing non-synchronous swimming activities (Santos et al., 2003, Pharmacol Biochem Behav 75:25-33). Conversely, although the preferential H(3)R antagonist (thioperamide) did not interfere with normal swimming behaviors, it surprisingly did ameliorate heavy metal-dependent hyperactive states (Giusi et al., 2008, Toxicol Appl Pharmacol 227:248-256). In the case of the xenoestrogens atrazine and endosulfan, their actions tend to mostly account for feeding alterations through hypothalamic H(3)R-dependent mechanisms. The aim of this review is to highlight the type of ED-HAergic neuroreceptor variations that are involved in stressor-dependent neurobehavioral responses of commercially valuable marine teleosts.

Recent evidences indicate that cerebral neurotrophic factors like vascular endothelial growth factor plus signaling pathways of the glutamatergic neuroreceptor system (L-Glu) are determinant modulators of depression-like states. In the present study, the type of interaction(s) exerted by the AMPAergic antagonist, 6-cyano-7-nitroquinoxalin-2,3-dione (CNQX) and the brain derived neurotrophic factor (BDNF) on depression-like behaviors in hamsters (Mesocricetus auratus) were investigated. Sub-chronic administration of BDNF in the hippocampal dentate gyrus (DG) of stressed hamsters was responsible for very evident (p

In this work, we describe the development of a compartmentalized membrane system using neonatal rodent hippocampal cells and human mesenchymal stem cells (hMSCs) to investigate the neuroprotective effects of hMSCs. To elucidate this interaction an in vitro oxygen-glucose deprivation (OGD) model was used that mimics central nervous system insults in vivo . Cells were cultured in a membrane system with a sandwich configuration in which the hippocampal cells were seeded on a fluorocarbon (FC) membrane, and were separated by hMSCs through a semipermeable polyethersulfone (PES) membrane that ensures the transport of molecules and paracrine factors, but prevents cell-to-cell contact. This system was used to simulate a cerebral ischemic damage by inducing OGD for 120 min. The core contribution of the work highlights the neuroprotective effects of hMSCs on hippocampal cells in a membrane system for the first time. The novel results show that hMSC secretome factors protect hippocampal cells against OGD insults as indicated by the conservation of specific structural and functional cell features together with the development of a highly branched neural network after the damage. Moreover, neuronal cells co-cultured with hMSCs before OGD insult were able to maintain BDNF production and O 2 consumption and did not express the apoptotic markers that were expressed in similarly insulted neuronal cells that had not been co-cultured with hMSCs. This compartmentalized membrane system appears to be a very useful and reliable system for studying the neuroprotective effects of hMSCs and identifying secreted factors that may be involved. Statement of Significance This paper is based on a combined synergism of biomaterials technology and stem cell approach, focusing on the development of a compartmentalized membrane system that serves as an innovative tool for highlighting the role of hMSCs on hippocampal neurons upon damage. The membrane system consists of two different flat sheet membranes, giving rise to double and separated cell membrane compartments that prevent cell-to-cell contact but allow the transport of paracrine factors. This system strongly corroborates the paracrine mediated neuroprotection of hMSCs on ischemic damaged neurons. The challenging and pioneeristic approach by using biomaterials allowed to perform a stepwise analysis of the phenomena, providing new insights into the field of MSC therapy.

Brain ischemia is a major cause of death and the most common element of disability in the adult population worldwide. This phenomenon occurs when blood flow is reduced or interrupted in the various brain districts leading to oxygen and glucose deprivation (OGD), which by triggering a complex series of biochemical plus molecular mechanisms such as an exacerbated production of misfolded and oxidized proteins together with the breakdown of cellular integrity are responsible for neuronal cell death. Despite the mechanisms that underlie the triggering of ischemic insults are still unclear, numerous studies are pointing to excitotoxic glutamatergic neuronal signaling processes as key mediators of these events. Indeed, cultured neurons deriving from global cerebral ischemia appear to respond to OGD with a rapid internalization of α ‑ amino ‑ 3 ‑ hydroxy ‑ 5 ‑ methyl ‑ 4 ‑ isoxazolepropionic acid receptors (AMPAR) thereby suggesting them as critical components of OGD-induced cell death. It strongly seems that OGD-dependent neuronal ischemia occurs via GluR2-sites in which a switching from GluR2-containing Ca 2+ -impermeable receptors to GluR2-lacking Ca 2+ -permeable subtypes constitutes an important step. Interestingly attention regarding excitotoxicity-related ischemic events, aside being largely directed to the o veractivation of AMPARs, appear to be also tightly linked to the translocation of the pro-apoptotic protein Bax to the mitochondria accounting for the activation of the caspase factors. Although the brain is able to repair part of the neuronal damages and to restore the morpho-functional organization, cerebral ischemia more than ever continues to attract much attention especially due to its elevated mortality feature. In this review we analyzed the role played by GluR2 AMPAR subunit in the pathological processes that lead to neurodegenerative diseases with particular attention being paid to the assembly of the major synaptic AMPARs together with cellular events that feasibly account for ischemic brain damages. In this context, knowledge of the different molecular mechanisms operating under these conditions may surely provide helpful indications regarding the identification of new therapeutic targets for treating cerebral ischemia.

The hibernating golden hamster (Mesocricetus auratus) is becoming a useful rodent model to study the neurophysiological role of some neuromediators on vital behaviors such as sleep and thermoregulation. Recent works have shown that the histamine neuroreceptor subtypes (H-sub(1-3)R) are able to modulate such behaviors. Here, specific subtype(s) and cerebral nuclei that were actively operating on feeding behaviors in pubertal and adult hamsters were identified. Of the subtypes assessed, H-sub-3R antagonist (thioperamide) provoked significant (p < .001) changes in behavior (very low total food and water intake) in adults, whereas it did not significantly modify these behaviors in pubertals. The H-sub-3R antagonist's role seemed to be related to elevated amounts of stress-induced damaged neurons displaying, mainly, shrunken crenated cell membranes and altered synaptic processes in limbic areas such as amygdala, cortex, and hippocampus. At the transcription level, an evident expression pattern of H-sub-3R messenger RNA appeared in pubertals, especially in neurons of the cortex and hippocampus, whereas the same trend was featured in amygdalar areas of hibernating adult hamsters, suggesting early H-sub-3R regulatory activities, at least in limbic sites of this rodent model.

It has now been established that a large number of man-made and natural chemicals are capable of interfering with the action of natural hormones. In this category "endocrine disruptors" such as the herbicide atrazine, when administered at ecological low doses (1 or 100 microg/kg per day) from gestational day 14 to postnatal day 21, provided a clear dimorphic neurodegenerative pattern in some brain areas of the domestic mouse (Mus musculus). Indeed, the high concentration (100 microg/kg per day) with respect to the low concentration (1 microg/kg per day) induced relevant neuronal damage in extrahypothalamic sites, such as the cortical and striatal areas in both sexes. Marked alterations in other areas, including the hippocampal and hypothalamic nuclei, were mostly typical of the female. At the neuronal level, the neuropeptide somatostatin, specific for the secretion of growth hormone, seemed to be a major target of atrazine effects, as demonstrated by evident subtype2,3,5 receptor mRNA differences of this neuropeptide, at least for the first two subtypes. In particular, a very strong (p0.001) upregulation of subtype2 expressing neurons was detected in female hypothalamic areas, specifically the suprachiasmatic nucleus, whereas a similar downregulatory trend was reported for some extrahypothalamic areas such as the striatum. Interestingly, very strong upregulatory and downregulatory actions were detected for neurons expressing subtype3 in male hypothalamic and amygdalar regions and in the cortical and hippocampal areas, respectively. Overall, it appears that these first neurotoxicological effects of atrazine are very likely linked to dimorphic expression patterns of specific somatostatin subtypes in discrete but key hypothalamic and extrahypothalamic areas of Mus musculus.

Common environmental contaminants such as heavy metals and pesticides pose serious risks to behavioral and neuroendocrine functions of many aquatic organisms. In the present study, we show that the heavy metal cadmium and the pesticide endosulfan produce such effects through an interaction of specific cerebral histamine receptor subtypes in the teleost ornate wrasse (Thalassoma pavo). Treatment of this teleost with toxic cadmium levels for 1 week was sufficient to induce abnormal swimming movements, whereas reduced feeding behaviors were provoked predominantly by elevated endosulfan concentrations. In the brain, these environmental contaminants caused neuronal degeneration in cerebral targets such as the mesencephalon and hypothalamus, damage that appeared to correlate with altered binding levels of the three major histamine receptors (subtypes 1, 2, and 3). Although cadmium accounted for reduced binding activity of all three subtypes in most brain regions, it was subtype 2 that seemed to be its main target, as shown by a very great (p < 0.001) down-regulation in mesencephalic areas such as the stratum griseum central layer. Conversely, endosulfan provided very great and great (p < 0.01) up-regulating effects of subtype 3 and 1 levels, respectively, in preoptic-hypothalamic areas such as the medial part of the lateral tuberal nucleus, and in the suprachiasmatic nucleus. These results suggest that the neurotoxicant-dependent abnormal motor and feeding behaviors may well be tightly linked to binding activities of distinct histamine subtypes in localized brain regions of the Thalassoma pavo.

The neurotoxic 3-nitropropionic acid (3-NP), a freckled milk vetch-derived inhibitor of mitochondrial enzymatic processes that is capable of mimicking the typical pathological features of neurodegenerative disorders, behaved in a differentiated manner in a hibernating rodent (hamster) with respect to a nonhibernating rodent (rat). Treatment of the two rodents with both an acute and chronic 3-NP dose supplied deleterious neuronal effects due to distinct histamine receptor (H(n)R) transcriptional activities, especially in the case of the rat. In hamsters, these treatment modalities accounted for overall reduced global activity in a freely moving environment and overt motor symptoms such as hindlimb dystonia and clasping with respect to the greater abnormal motor behaviors in rats. This behavioral difference appeared to be strongly related to qualitative fewer neuronal alterations and, namely, lesser crenated cell membranes, swollen mitochondria, and darkened nuclei in hamster brain areas. Moreover, a mixed H(1,3)R mRNA expression pattern was reported for both rodents treated with a chronic 3-NP dose as demonstrated by predominantly low H1R mRNA levels (>50%) in rat striatum and cortex, whereas extremely high H3R levels (>80%) characterized the lateral and central amygdala nuclei plus the striatum of hamsters. Interestingly, the H3R antagonist (thioperamide) blocked 3-NP-dependent behaviors plus induced an up-regulation of H1R levels in mainly the above-reported hamster amygdalar nuclei. Overall, these results show, for the first time, that a major protective role against neurodegenerative events appears to be strongly related to the expression activity of H(1,3)R subtypes of amygdalar neurons in this hibernating model.

Characterization of somatostatinergic (sst) neuronal activity through the application of nonpeptidyl agonists L-779,976 and L-817,818 which are highly specific for the sst receptors (sstr) sstr2 and sstr5, respectively, shows for the first time that sstr2, 5-like subtypes are the two major sstr subtypes operating in the brain of the teleost sea wrasse, Coris julis. A somewhat high but heterogeneous distribution pattern (>30

In an effort to establish distinct phylogenetic values for the major GABA(A) receptor subunit mRNAs (alpha(1), beta(2) and gamma(2)) in the telencephalon, the cellular expression capacity of these mRNAs was evaluated in this brain region of two distantly related mammals (hedgehog and rat). Application of emulsion hybridization histochemistry and quantitative film autoradiography approaches allowed us to detect elevated ( P0.01) beta(2) mRNA levels in primitive telencephalic neuronal populations such as those of the glomerular olfactory layer of the hedgehog. Conversely, notable levels of the alpha(1)-subunit mRNA were also registered for the higher telencephalic regions such as the oriens-pyramidalis layer of the hippocampus although elevated levels were mostly obtained for the rat. Interestingly, it was the high number of giant pyramidal neurons that were actively involved in the expression of this subunit. A similar pattern was also detected for pyramidal-like cells of the rat lateral amygdala region expressing the gamma(2)-subunit mRNA. Taken together, these results suggest an early phylogenetic success of GABAergic neuronal mechanisms relying mainly on elevated beta(2)-subunit levels. On the other hand, elevated quantities of gamma(2)- and, above all, of alpha(1)-subunit mRNAs in the rat seem instead to be related to the highly advanced behavioral and mnemonic strategies that are under the control of these telencephalic regions.

Hibernation is a physiological state that by putting vital biological processes at rest enables mammals to protect all organs, especially the brain against ischemic insults and reperfusion injuries. Earlier studies have highlighted the role of hypothalamic (HTH) sites like the periventricular nucleus (Pe) toward sleep-wake and cardiovascular activities of hibernators. In the present work, infusions of Pe with the orexigenic neuropeptide orexin-A (ORX-A) or the novel anti-obesity sympathoinhibitory neuroactive peptide catestatin (CST) have been correlated to differing feeding and motor behaviors in the facultative hibernating hamster Mesocricetus auratus. Behavioral observations showed that treatment with CST provided an anti-obesity activity via the reduction of food intake and body weight for all hibernating states, while ORX-A promoted orexigenic events during mainly the entrance phase. Moreover, hamsters treated with this neuropeptide during the entrance and the arousal hypertensive phases also featured elevated ORX 2 receptor (ORX2R) levels in the third layer of the parietal cortex and lateral HTH (LH), areas involved with feeding, motor plus sleep-wake rhythms. Conversely, ORX-A down-regulated ORX2Rs in the ventromedial (VMH) and supraoptic (SO) HTH nuclei that are associated with anorexigenic effects. Even CST induced mixed ORX2R expression patterns in mostly HTH areas like the evident down-regulation in LH along with the up-regulation in VMH and SO. Overall treatments, especially ORX-A+CST led to reduced neurodegenerative phenomena in HTH supporting their importance together with ORX2Rs in preserving hemodynamic activities, feeding and sleep-wake rhythms of this diencephalic station, which may supply useful therapeutic indications for treating cardiovascular disturbances linked with brain dysfunctions.

The relationship of morphometrical and androgen receptor evaluations of the main testicular interstitium cellular element (Leydig cells) in the domestic pig provided interesting numerical and morphological features during the different aging stages. As early as 25 days (a period in which the pig is sexually immature) there was a low number of Leydig cells (1.46 x 10(8)) with respect to a 78% and 35% increase in the adult (2.48 x 108) and aged (1.78 x 10(8)) animal, respectively. Interestingly, when the volume density of Leydig cells was considered, the average volume of these cells seemed to be high (75%) in the aged pig with respect to the young immature animal whereas a lower increase (27%) was observed for the adult animal. Moreover, the evaluation of testosterone receptor binding sites in the testis at the various stages of development also displayed a differentiated pattern since elevated testosterone receptor binding levels of the high dissociation affinity type were obtained for the adult pig. Thus, from the combined morphological variations of Leydig cells and testosterone receptor binding activity, it appears that this androgenic receptor component exerts distinct autocrine effects on the different functional features of some testicular tissue constituents at the different aging stages of the domestic pig.

This investigation represents a first study dealing with the dimorphic differences of the main alpha-containing gamma-aminobutyric acid (GABA(A)) receptors in the brain of two distantly related mammals (hedgehog and rat). The labeling of these receptors in the presence of zolpidem (highly selective benzodiazepine agonist) and under the different degree of GABA(A)/benzodiazepine allosteric coupling activities accounted for a heterogeneous colocalization of alpha1-enriched and of alpha2/3-enriched and alpha5-enriched GABA(A) receptors in some areas of the cortico-basal ganglia system (including the important ventrolateral thalamic station) of both mammalian sexes. In the hedgehog, the greatest (P0.001) GABA-dependent reduction of zolpidem inhibition constants was mostly registered in alpha1- and/or alpha5-enriched areas, such as the frontoparietal cortex lamina III (235%), ventrolateral thalamic nucleus (128%), and substantia nigra pars reticulata (110%) of the male. However, the greatest reductions in the rat were instead detected in the male substantia nigra pars reticulata (192%) and female striatum (120%), areas which are enriched either by the colocalization of alpha1- with alpha2/3-subunits or by all three alpha-subunits. These results support the contention that a sex-related alpha-containing GABA(A) receptor sensitivity constitutes an important element in the execution of skilled motor activities during the different socio-sexual behaviors of the two mammals.

Different binding activities of 125I-rat atrial natriuretic peptide were evaluated using in vitro quantitative autoradiography in the heart of two antarctic notothenioid teleosts, the red-blooded Trematomus bernacchii and hemoglobinless Chionodraco hamatus. Saturable and specific binding sites for rat atrial natriuretic peptide were found in the atrium, ventricular myocardium, ventricular endocardium and inner and outer layers of the bulbus arteriosus of both fishes. Scatchard analysis of the saturation data showed that the atrium, ventricular endocardium and outer bulbar layer of T. bernacchii were characterized by a single class of high affinity natriuretic peptide binding sites (Kd = 14 ± 3.2, 9.7 ± 2.3 and 6.2 ± 1.3 pM, respectively), whereas the ventricular myocardium and the inner bulbar layer contained elevated numbers of two classes of high and low affinity natriuretic peptide binding sites (1.8 ± 0.6 < Kd < 209 ± 66 pM). In contrast, in C. hamatus, both high and low affinity binding sites were detected in all cardiac regions (2.1 ± 0.7 < Kd < 262 ± 90 pM). In both fishes, competition experiments in the presence of either unlabeled rat atrial natriuretic peptide or porcine brain natriuretic peptide indicated different displacement capacities. Porcine brain natriuretic peptide, able to bind to natriuretic peptides sites in all the heart regions of both notothenioids, provided a higher displacement capacity with respect to that of rat atrial natriuretic peptide in the atrium of T. bernacchii.

Fish are particularly sensitive to copper (Cu) because although it is an essential metal, it becomes dangerous for aquatic ecosystems, thus accounting for physiological alterations. In this study, we investigated Cu effects on neurobehavioral activities of Thalassoma pavo and, above all, its recovery strategies by evaluating behavioral disturbances, neurodegeneration, and expression of heat shock protein (HSP)90 and orexin receptor (ORXR). Ornate wrasses exposed to nominal sublethal higher (1.07 mg/l) and lower (0.25 mg/l) concentrations of CuCl₂·H₂O mostly reduced swimming and feeding activities along with inducing abnormal behaviors in a dose-dependent manner. Concomitantly, ORXR transcripts were mostly upregulated in the diffuse nucleus of the inferior lobe (NDLI, + 142%) and the corpus of the cerebellum (CCe, + 243%), whereas HSP90 was downregulated only in lateral part of the dorsal telencephalon (Dl, -35%), the nucleus glomerulosus (NG; -40%), and the optic tectum (OT; -33%). Interestingly, when fish were transferred to Cu-free water, some behaviors were promptly rescued, especially in fish previously exposed to the lower Cu concentration. This rescuing tendency was confirmed by evident reductions of argyrophilic signals in Dl (-67%), NG (-31%), and OT (-42%) of fish exposed to 0.25 mg/l Cu. Moreover, transcriptional events of both ORXR and HSP90 were further upregulated in order to orchestrate a reactivation of behavioral and neuronal functions. Overall, this study highlights, for the first time, new neuronal strategies against environmental adverse conditions involving both ORXergic system and HSP90 as key elements that may assure, at least in part, protection and recovery processes against toxic agents.

Recent interferences of hemodynamic functions via modified brain neuronal mechanisms have proven to be major causes of dementia and sleeping disorders. In this work, cerebral expression differences of the neuroactive vesicular chromogranin A (CgA) and distinct α GABAAR subunits were detected in the facultative hibernating hamster. In particular, damaged neuronal fields of hypotensive torpor (TORP) state were correlated to elevated CgA and GABAAR α1, α4 mRNA levels in the paraventricular hypothalamic nucleus (PVN), central amygdalar nucleus (CeA) plus solitary tractus nucleus (NTS). Conversely, few neurodegeneration signals of hypertensive arousal (AROU) state, accounted for mostly lower CgA levels in the same areas. This state also provided increased α2-containing sites in amygdala, hippocampal and NTS neurons together with elevated α4-containing receptors in the periventricular hypothalamic nucleus (Pe). Interestingly in our hibernating model, CgA appeared to preferentially feature inhibitory neurosignals as indicated by preliminary perfusion of amygdalar sites with its highly specific antihypertensive derived peptide (catestatin) promoting GABA-dependent sIPSCs. Overall, evident neuronal damages plus altered expression capacities of CgA and α1-, α2-, α4-GABAARs in CeA, Pe, PVN as well as NTS during both hibernating states corroborate for the first time key molecular switching events guaranteeing useful cardiovascular rescuing abilities of neurodegenerative disorders.

A distinct distributional pattern of natriuretic peptide receptors has been demonstrated by in vitro quantitative autoradiography in the heart of Anguilla anguilla where saturable, stable, and specific binding sites for rat atrial natriuretic peptide in atrium, ventricle, and bulbus arteriosus have been detected. Competition experiments in the presence of either rat atrial natriuretic peptide or unlabeled eel peptides, i.e., atrial natriuretic peptide, C-type natriuretic peptide, and ventricular natriuretic peptide, showed differences in the relative affinity orders of these peptides in the various heart chambers. In the atrium and in the ventricular myocardium most receptors likely are type C, while B-type receptors are present in significant amounts in the ventricular endocardium and in the inner and outer layers of the bulbus arteriosus. Scatchard analysis of the competitive data indicates that in the atrium and in the ventricular myocardium only one class of receptors for rat atrial natriuretic peptide is present (Kd = 105 ± 16 and 107 ± 15 pM, respectively), while the ventricular endocardium and the inner and outer layers of the bulbus arteriosus contain two classes of binding sites (6 ± 3.7 < Kd < 20 ± 3.9 pM for site 1; 148 ± 72 < Kd < 271 ± 52 pM for site 2). © 1996 Wiley-Liss, Inc.

The hindering effects of metals and in particular lead (Pb) are representing a growing threat to aquatic organisms such as fish. This observation derives from toxic concentrations of Pb accounting for altered neurophysiological activities of some interesting teleost models like Thalassoma pavo, a fish species highly known for its host-cleaning symbiosis. In this study, the nominal PbNO(3) concentration of 1.6 mg/L was capable of reducing feeding and resting bouts as early as 24 h of exposure while hyperactive swimming episodes were also detected. Such abnormal behaviors were tightly correlated to up-regulated orexin receptor (ORXR) mRNA expression levels in some brain areas such as the lateral thalamic nucleus (+213%) and the optic tectum (+90%) with respect to controls. Interestingly, these transcriptional effects seemed to be attenuated when Pb-exposed fish received either 100 ng/g of ORX-A (-70%) or 0.1 μg/g of γ-aminobutyric acid(A) receptor (GABA(A)R) agonist muscimol (MUS; -97%) compared to fish exposed to Pb alone. Moreover, a net neurodegenerative process of the different brain areas was reported after Pb exposure as displayed by their marked amino cupric silver stained cells while these cells were devoid of any staining reaction after treatment with MUS only. Conversely, addition of the GABA(A)R antagonist bicuculline (BIC; 1 μg/g) moderately (p

Recently, environmental stimuli on different neurobiological events, via participation of distinct amygdalar (AMY) ORXergic fibers have aroused wide interests in view of their ability to modify neuronal linked stressful and physiological homeostatic conditions. Results of the present study indicate that ORXergic (ORX-A/B) circuits of the facultative hibernating golden hamster (Mesocricetus auratus) central AMY (CeA) and basolateral AMY (BlA) nuclei constitute major sites of feeding behaviors. Indeed, hamsters after treatment of BlA with ORX-A frequently ingested greater quantities of food as compared to controls, while ORX-B in CeA induced a very (p0.001) great consumption of water. The same nuclei treated separately with either ORX-A or ORX-B ± the selective α(1) GABA(A) benzodiazepine receptor agonist (zolpidem) dedicated less time to eating and drinking sessions. Conversely, hamsters that received the same neuropeptides but this time with the glutamatergic agonist NMDA displayed greater hyperphagic effects above all for ORX-A. When behavioral changes were compared to the expression of the specific ORXergic receptor (ORX-2R), an up-/down-regulating pattern was detected in some limbic areas (AMY, hippocampus and hypothalamus) following treatment with ORX-A or ORX-B plus NMDA. Overall, indications deriving from this study strongly point to hamster BlA-enriched ORX-A fibers in combination with either inhibitory or excitatory signals as main targets of hyperphagic responses while CeA ORX-B activities in presence of these same neuronal signals predominantly induced drinking motivational behaviors. The distinct behavioral activities of these two neuropeptides may have useful clinical bearings toward psychiatric and sleeping disorders such as bulimia and narcolepsy.

Background The availability of oxygen is a limiting factor for neuronal survival since low levels account not only for the impairment of physiological activities such as sleep-wake cycle, but above all for ischemic-like neurodegenerative disorders. In an attempt to improve our knowledge concerning the type of molecular mechanisms operating during stressful states like those of hypoxic conditions, attention was focused on eventual transcriptional alterations of some key AMPAergic silent neuronal receptor subtypes (GluR1 and GluR2) along with HSPs and HIF-1α during either a normoxic or a hypoxic aestivation of a typical aquatic aestivator, i.e. the lungfish (Protopterus annectens). Results The identification of partial nucleotide fragments codifying for both AMPA receptor subtypes in Protopterus annectens displayed a putative high degree of similarity to that of not only fish but also to those of amphibians, birds and mammals. qPCR and in situ hybridization supplied a very high (p p Conclusions The distinct transcriptional variations of silent neurons expressing GluR1/2 and HSPs tend to corroborate these factors as determining elements for the physiological success of normoxic and hypoxic aestivation. A distinct switching among these AMPA receptor subtypes during aestivation highlights new potential adaptive strategies operating in key brain regions of the lungfish in relation to oxygen availability. This functional relationship might have therapeutic bearings for hypoxia-related dysfunctions, above all in view of recently identified silent neuron-dependent motor activity ameliorations in mammals.

To date, many scientific attempts have been directed towards the development of experimental models for the identification of neuronal mechanisms evoking cardiovascular and hemodynamic dysfunctions. The spontaneously hypertensive rat (SHR), a genetic model of essential hypertension, has become a valuable rodent for the characterization of molecular markers in hypertensive-related diseases. Recently, growing interests have also been directed to a new experimental paradigm i.e. hibernation, a physiological state consenting the hamster (Mesocricetus auratus) to activate protective mechanisms against ischemic-like complications during the arousal phase. With this intention, the present review will focus attention on specific neurosignaling systems involved with the preservation of hemodynamic conditions in those brain areas that play a pivotal role on such a feature. It is widely known that healthy neurons conserve their structural and responsiveness properties in presence of a constant blood supply, which is assured by their coupling to microvessels and perivascular astrocytes as well as by secretory proteins such as chromogranin A (CgA). So, it will be interesting to establish if this protein alone or with the participation of excitatory/inhibitory neurosignals is capable of influencing some brain areas controlling cardiovascular conditions in both SHRs and hibernating hamsters. In this context, the present work will deliver the most important findings regarding neuronal CgA and its cross-talking ability with major inhibitory (GABA/adenosine) and/or excitatory (glutamate) neuroreceptor systems in relation to hypertensive/hypotensive states of both animal models. Indications deriving from such approaches may provide clinically useful insights regarding their role as protective factors of hemodynamic and neurological disorders.

Quantitative receptor autoradiography was used to study the binding of 2-[125I] iodomelatonin in the brain of the castrated and gonadally intact male Japanese quail Coturnix japonica exposed to both long- and short-day photoperiod cycles. The distribution study displayed that these conditions were responsible for a heterogeneous binding pattern as shown by elevated receptor levels being located in visual brain centers, such as the stratum opticum, nucleus pretectalis, and nucleus geniculatus lateralis, pars ventralis, while lower values were found in the nucleus lateralis hypothalami and nucleus isthmi pars magnocellularis. Closer examination of the 2-[125I] iodomelatonin–binding pattern following the different gonadal and photic influences showed that combination of the gonadally intact condition and a 16L:8D (long-day) photoperiod cycle was required for the greater binding changes. These differences occurred in brain sites such as the area preoptica, paleostriatum primitivum, and nucleus ectomamillaris. Saturation binding studies, which were carried out only in some of the above areas, revealed that the combined gonadal and photic-induced changes are basically due to the modifications of total number of binding sites. The importance of a gonadal steroid modulatory role in the photic-dependent melatonin binding activity suggests that other types of neuronal mechanisms might be involved in the regulation of neuroendocrine and socio–sexual behaviors in nonmammalian vertebrates. © 1994 Wiley-Liss, Inc.

Orexins are one of the most potent orexigenic factors in fish that through their interaction with the GABA(A) receptor system assures the successful execution of feeding, motor and sleep-wake activities. In the present study, the effects of ORX-A (10ng/g BW) very greatly enhanced (p

Background Excitatory transmitting mechanisms are proving to play a critical role on neuronal homeostasis conditions of facultative hibernators such as the Syrian golden hamster. Indeed works have shown that the glutamatergic system of the main olfactory brain station (amygdala) is capable of controlling thermoregulatory responses, which are considered vital for the different hibernating states. In the present study the role of amygdalar glutamatergic circuits on non-hibernating (NHIB) and hibernating (HIB) hamsters were assessed on drinking stimuli and subsequently compared to expression variations of some glutamatergic subtype mRNA levels in limbic areas. For this study the two major glutamatergic antagonists and namely that of N-methyl-D-aspartate receptor (NMDAR), 3-(+)-2-carboxypiperazin-4-yl-propyl-1-phosphonate (CPP) plus that of the acid α-amine-3-hydroxy-5-metil-4-isoxazol-propionic receptor (AMPAR) site, cyano-7-nitro-quinoxaline-2,3-dione (CNQX) were infused into the basolateral amygdala nucleus. Attempts were made to establish the type of effects evoked by amygdalar glutamatergic cross-talking processes during drinking stimuli, a response that may corroborate their major role at least during some stages of this physiological activity in hibernators. Results From the behavioral results it appears that the two glutamatergic compounds exerted distinct effects. In the first case local infusion of basolateral complexes (BLA) with NMDAR antagonist caused very great (p < 0.001) drinking rhythms while moderately increased feeding (p < 0.05) responses during arousal with respect to moderately increased drinking levels in euthermics. Conversely, treatment with CNQX did not modify drinking rhythms and so animals spent more time executing exploratory behaviors. These same antagonists accounted for altered glutamatergic transcription activities as displayed by greatly reduced GluR1, NR1 and GluR2 levels in hippocampus, ventromedial hypothalamic nucleus (VMN) and amygdala, respectively, plus a great (p < 0.01) up-regulation of GluR2 in VMN of hibernators. Conclusion We conclude that predominant drinking events evoked by glutamatergic mechanisms, in the presence of prevalently down regulated levels of NR1/2A of some telencephalic and hypothalamic areas appear to constitute an important neuronal switch at least during arousal stage of hibernation. The establishment of the type of glutamatergic subtypes that are linked to successful hibernating states, via drinking stimuli, may have useful bearings toward sleeping disorders.

The hippocampus is recognized as a major telencephalic area modulating learning and episodic memory through the activation of its different subregions. The various functional properties of Ammon's horn 1 (Cornu Amonis 1; CA1) area have been shown to rely on GABAergic and Glutamat- (Glu)-ergic neuronal signals during both postnatal and adult stages. For this purpose, it was the aim of the present study to establish whether certain alpha GABA(A)R subunits (alpha(2,5)) were capable of modifying CA1 structural and functional features via their interaction with specific NMDA receptor subunits such as NR1 during early development stages of the hibernating hamster (Mesocricetus auratus). Indeed, in vitro addition of the selective alpha(2,5) GABA(A)R agonist diazepam (DZP; alpha(2,5)) accounted for early neuronal formations that were blocked by its antagonist flumazenil (FLM). In particular, the former drug caused very great (p0.001) increases of dendritic sprouting and branching processes mainly at day in vitro (DIV) 3, while its effects still continued to be responsible for moderate (p0.05) increases of axonal length during the entire culture period. Contextually, DZP was also responsible for a very great up-regulated expression of neuritic NR1 and MAP2 together with a great (p0.01) increase of synaptophysin at DIV7. Overall, this first study suggests a specifically tight cross-talking relationship of GABAergic/Gluergic mechanisms operating during CA1 neuronal development, which may bring us closer to the identification of more selective therapeutic targets for hippocampal-linked neurological disorders.

The inhibitory effect of female sex steroid hormones on the binding of [35S]t-butylbicyclophosphorothionate [35S]TBPS to the chloride ion channel receptor in different forebrain areas of the female rat proved to be of a differential nature. The in vivo administration of estradiol and estradiol+progesterone were responsible for substantially lower chloride ion channel receptor levels in brain areas that contain elevated steroid receptors, such as the medial preoptic area, the cortico-medial amygdala nucleus, the vertical limb diagonal band-medial septal nucleus and the cortex lamina V. The administration of progesterone alone reduced receptor levels in the oriens-pyramidalis CA1 layer of the hippocampus, caudate putamen, cortex lamina VI (brain areas that contain little if any steroid receptors) and in the lateral and basolateral amygdala nucleus (brain sites that contain noninducible progesterone receptors). On the basis of the progesterone-inhibitory activity on the chloride ion channel receptors, it was important to investigate whether progesterone per se or whether the potent progesterone metabolites 3 alpha-hydroxy-5 alpha-dihydroprogesterone (3 alpha, 5 alpha-THP) and 3 beta-hydroxy-5 beta-dihydroprogesterone (3 beta,5 beta-THP) were involved in the binding level changes, and to establish the specific brain sites where these steroid effects occur. In fact, in vitro addition of the 5 alpha-reduced progesterone metabolite produced even greater depressive effects on [35S]TBPS binding not only in the same brain areas as the in vivo progesterone replacement therapy but also in some sites that provided significant receptor level changes following the sequential administration of estradiol+progesterone. However, when the 5 beta-reduced metabolite was tested on the binding of [35S]TBPS to the chloride ion channel receptor, only the basolateral amygdala nucleus, the cortex lamina VI and the dorsolateral septal nucleus exhibited changes. Because the steroid-mediated chloride ion flux is regulated in a GABA-dependent manner, we next checked for the type of GABA effects on the chloride ion channel receptor levels and found that GABA not only intensified the 3 alpha,5 alpha-THP inhibitory effects but, together with this progesterone metabolite, was also involved in binding changes in the vertical limb diagonal band-medial septal nucleus. It is interesting to note that the GABA effects on 5 beta-metabolite-induced receptor changes were not of the enhancing type, but tended, rather, to be inhibitory.(ABSTRACT TRUNCATED AT 400 WORDS)

Specific binding sites for GABAA and benzodiazepines were detected in the brain of the frog Rana esculenta after the in vitro incubation of tissue sections with their res

Decreased body temperature during hibernation evokes a neuroprotective effect against the frequent neurodegenerative events of ischemic/reperfusion injuries. This neuroprotection appears to stem from a direct involvement of orexin-A plus the sympathoinhibitory neuroactive peptide catestatin on orexin 2 receptor-dependent feeding and motor behaviors of the facultative hibernating hamster.

Ammonia in dipnoans plays a crucial role on neuronal homeostasis, especially for those brain areas that maintain torpor and awakening states in equilibrium. In the present study, specific α subunits of the major neuroreceptor inhibitory complex (GABAAR), which predominated during some phases of aestivation of the lungfish Protopterus annectens, turned out to be key adaptive factors of this species. From the isolation, for the first time, of the encoding sequence for GABAAR α1, α4, and α5 subunits in Protopterus annectens, qPCR and in situ hybridization levels of α4 transcript in thalamic (P < 0.001) and mesencephalic (P < 0.01) areas proved to be significantly higher during long aestivating maintenance states. Very evident α5 mRNA levels were detected in diencephalon during short inductive aestivating states, whereas an α4/α1 turnover characterized the arousal state. Contextually, the recovery of physiological activities appeared to be tightly related to an evident up-regulation of α1 transcripts in telencephalic and cerebellar sites. Surprisingly, TUNEL and amino cupric silver methods corroborated apoptotic and neurodegenerative cellular events, respectively, above all in telencephalon and cerebellum of lungfish exposed to long maintenance aestivating conditions. Overall, these results tend to underlie a novel GABAergic-related ON/OFF molecular switch operating during aestivation of the lungfish, which might have a bearing on sleeping disorders. © 2011 Wiley-Liss, Inc.

Background The structural arrangement of the γ-aminobutyric acid type A receptor (GABAAR) is known to be crucial for the maintenance of cerebral-dependent homeostatic mechanisms during the promotion of highly adaptive neurophysiological events of the permissive hibernating rodent, i.e the Syrian golden hamster. In this study, in vitro quantitative autoradiography and in situ hybridization were assessed in major hypothalamic nuclei. Reverse Transcription Reaction-Polymerase chain reaction (RT-PCR) tests were performed for specific GABAAR receptor subunit gene primers synthases of non-hibernating (NHIB) and hibernating (HIB) hamsters. Attempts were made to identify the type of αβγ subunit combinations operating during the switching ON/OFF of neuronal activities in some hypothalamic nuclei of hibernators. Results Both autoradiography and molecular analysis supplied distinct expression patterns of all α subunits considered as shown by a strong (p < 0.01) prevalence of α1 ratio (over total α subunits considered in the present study) in the medial preoptic area (MPOA) and arcuate nucleus (Arc) of NHIBs with respect to HIBs. At the same time α2 subunit levels proved to be typical of periventricular nucleus (Pe) and Arc of HIB, while strong α4 expression levels were detected during awakening state in the key circadian hypothalamic station, i.e. the suprachiasmatic nucleus (Sch; 60%). Regarding the other two subunits (β and γ), elevated β3 and γ3 mRNAs levels mostly characterized MPOA of HIBs, while prevalently elevated expression concentrations of the same subunits were also typical of Sch, even though this time during the awakening state. In the case of Arc, notably elevated levels were obtained for β3 and γ2 during hibernating conditions. Conclusion We conclude that different αβγ subunits are operating as major elements either at the onset of torpor or during induction of the arousal state in the Syrian golden hamster. The identification of a brain regional distribution pattern of distinct GABAAR subunit combinations may prove to be very useful for highlighting GABAergic mechanisms functioning at least during the different physiological states of hibernators and this may have interesting therapeutic bearings on neurological sleeping disorders.

Quantitative receptor autoradiographic study of 125I-atrial natriuretic peptide factor (ANF) in the heart of a teleost fish Conger conger has shown that a heterogenous distribution of 125I-ANF binding exists in the different cardiac regions. Elevated ANF binding densities (3,790 fmol/mg protein) were encountered in the innermost layer (tunica intima) of the bulbus arteriosus while lower binding levels (293–403 fmol/mg protein) were revealed in atrium and ventricle. In order to determine 125I-ANF binding characteristics (KD, Bmax) in the above cardiac sites, saturation binding assays were carried out. The results show that low 125I-ANF KD values (28.8–52.6 pM) were found in the atrium and in the bulbus arteriosus with respect to the higher KD values (373 pM) of the ventricle. The number of binding sites were respectively 632 and 1,279 fmol/mg protein for the atrium and the ventricle, while a substantially elevated Bmax of 7,235 fmol/mg protein was found for the bulbus arteriosus. These results may furnish some insights concerning ANF receptor binding activity and its putative regulatory role of different cardiac functions. © 1992 Wiley-Liss, Inc.

Quantitative receptor autoradiography was used to study the neuroanatomical distribution and effects of gonadal hormones on [3H] flunitrazepam binding in the male Japanese quail brain. In gonadally intact quail brains, [3H] flunitrazepam displayed a heterogeneous distribution, with elevated levels in the posterior brain regions such as the stratum griseum et fibrosum superficiale and stratum griseum centrale of the optic tectum. Lower values were observed in the anteriorly located brain sites such as the nucleus septalis (lateralis et medialis), the cortex dorsolateralis and the nucleus lateralis hypothalami. Castration affected [3H] flunitrazepam binding levels in brain areas known to contain gonadal steroid receptors as well as in some areas which were devoid of gonadal steroid receptors. Castration in fact, elevated [3H] flunitrazepam binding in the nucleus preopticus anterior and reduced binding levels in archistriatum dorsalis et ventralis and in the nucleus intercollicularis; all of these areas are known to have gonadal steroid receptors. In two regions which do not contain such receptors, namely the hyperstriatum ventrale and the cerebellum pars granularis, castration increased [3H] flunitrazepam binding. In order to determine whether the gonadal steroid effect is due to changes either in the number of binding sites (Bmax) and or affinity binding state (KD), saturation binding studies were carried out in some of the areas described above in brains of quail which were castrated or castrated and given replacement therapy with testosterone or estradiol for 2 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)

At date the major neuroreceptors i.e. gamma-aminobutyric acid(A) (GABA(A)R) and orexin (ORXR) systems are beginning to be linked to homeostasis, neuroendocrine and emotional states. In this study, intraperitoneal treatment of the marine teleost Thalassoma pavo with the highly selective GABA(A)R agonist (muscimol, MUS; 0.1 microg/g body weight) and/or its antagonist bicuculline (BIC; 1 microg/g body weight) have corroborated a GABA(A)ergic role on motor behaviors. In particular, MUS induced moderate (p

Picrotoxin blocking effects on GABA-induced inhibition of lateral merus display in the male crab Carcinus mediterraneus suggest that the chloride ion channel component of the GABA molecule is involved in the control of this defensive behavior. The low GABA doses (3 and 5 micrograms/g body weight) inhibited lateral merus display as early as 10 min after drug treatment while the administration of higher GABA concentrations (greater than 5 micrograms/g) prolonged the duration of the behavioral effects. The administration of 1 microgram/g picrotoxin to the animals treated with the effective GABA doses (5, 10 and 20 micrograms/g) restored lateral merus display. The antagonizing activity of picrotoxin on the inhibitory GABA effects, in a temporal manner, demonstrates that GABAergic sites other than peripheral ones are probably participating with the regulation of this agonistic posture. Quantitative autoradiography results revealed interesting receptor levels changes of the specific chloride ion channel antagonist [35S] t-butylbicyclophosphorothionate in the different brain areas of the male crab displaying lateral merus with respect to normal nonreactive animals. Elevated receptor binding levels were encountered in the middle and posterior brain area whereas low levels were obtained at the anterior level. Both the behavioral and autoradiographic results suggest that the defensive type of aggression behavior in crustacean may not only be mediated by a peripherally controlled GABA-gated chloride ion flux but also by a central GABAergic mechanism.

The neuropeptides hypocretins/orexins (ORX) are known to control state-dependent activities such as sleep-wakefulness, energy homeostasis, thermoregulation, and maternal behaviors. To date, interests regarding environmental-related ORX-ergic neuronal functions have dealt with mammals; only recently is attention beginning to be directed toward aquatic vertebrates. Here, photoperiod-dependent effects of ORX-A on behavioral, neurodegenerative and transcriptional activities were evaluated in some forebrain areas of a teleost Labridae (ornate wrasse, Thalassoma pavo). The ornate wrasse, when treated intraperitoneally with a high physiological dose (100 ng/g) of ORX-A and exposed to a natural photoperiod (12L:12D), exhibited very high (P0.001) locomotion and feeding behaviors. ORX-A in the presence of a constant light photoperiod accounted for numerically even greater (500%) feeding frequencies. Conversely, constant dark conditions very strongly reduced feeding habits and moderately (P0.05) increased resting states. In this case, the same ORX-A and photoperiodic conditions responsible for altered behaviors also induced neurodegenerative processes in the different hypothalamic, mesencephalic, and telencephalic neuronal fields. Interestingly, this ORX-A treatment seemed to be correlated to greater up-regulatory patterns of ORX receptor mRNA in these same brain areas, above all under constant light conditions rather than natural photoperiod. On the other hand, telencephalic sites provided a very active expression capacity during the dark phase. Overall, these results suggest for the first time that at least in the ornate wrasse, light- and dark-dependent ORX-ergic neuronal activities are able to cause short- and long-term abnormal motor behaviors, likely through neurodegenerative and transcriptional events in a brain regional manner.

The isolation of the G-protein-coupled receptor 30 (GPR30), an orphan membrane receptor unrelated to nuclear estrogen receptors (ERs), has become a key factor towards the unraveling of rapid estrogen action. This membrane receptor together with cellular signaling intermediaries, i.e., extracellular signal-dependent kinases 1 and 2, may promote neuronal proliferation and differentiation activities. In the present study, an evident gene expression pattern of GPR30 characterized postnatal 7 (young) and 60 (adult) days of age hamsters as shown by its heterogeneous mRNA distribution in hypothalamic, amygdalar and cerebellar areas of both sexes. In particular, most of the brain areas considered in the adult hamster plus only the amygdala and cerebellum of young animals behaved in a sexually dimorphic fashion. This similar pattern was also detected for the ERα and β, as shown by the latter receptor prevailing in young and adult females, while the former predominated in young females. Even for the two kinases, a sexually dimorphic distribution was featured above all for young hamsters. Overall, the findings of the present study established a distinct expression pattern of the novel ER (GPR30) that may operate differently in some brain areas of the hamster and this may provide interesting insights regarding its probable neuroprotective role during the execution of some hibernating states, which are typical of our rodent model.