Search

A population of Neuropeptide Y (NPY) neurons which co-express Agouti-related peptide (AgRP) in the arcuate nucleus of the hypothalamus (ARC) are inhibited at physiological levels of brain glucose and activated when glucose levels decline (e.g. glucose-inhibited or GI neurons). Fasting enhances the activation of NPY/AgRP-GI neurons by low glucose. In the present study we tested the hypothesis that lipopolysaccharide (LPS) inhibits the enhanced activation of NPY/AgRP-GI neurons by low glucose following a fast. Mice which express green fluorescent protein (GFP) on their NPY promoter were used to identify NPY/AgRP neurons. Fasting for 24 hours and LPS injection decreased blood glucose levels. As we have found previously, fasting increased c-fos expression in NPY/AgRP neurons and increased the activation of NPY/AgRP-GI neurons by decreased glucose. As we predicted, LPS blunted these effects of fasting at the 24 hour time point. Moreover, the inflammatory cytokine tumor necrosis factor alpha (TNFα) blocked the activation of NPY/AgRP-GI neurons by decreased glucose. These data suggest that LPS and TNFα may alter glucose and energy homeostasis, in part, due to changes in the glucose sensitivity of NPY/AgRP neurons. Interestingly, our findings also suggest that NPY/AgRP-GI neurons use a distinct mechanism to sense changes in extracellular glucose as compared to our previous studies of GI neurons in the adjacent ventromedial hypothalamic nucleus.

Moving visual stimuli elicit a sequence of coordinated activity of muscles including tongue protraction. Morphological and physiological studies fail to reveal any direct tectal projections to hypoglossal motoneurons suggesting that the last-order premotor interneurons (LOPI) are the direct recipients of neural activities generated in the optic tectum. The aim of this study is to analyze the topographical organization of the last-order premotor interneurons related to protractor muscles of the tongue. In Rana esculenta, biotinylated dextran amine (BDA) was injected by iontophoresis into the subnucleus of the hypoglossal nerve containing the motoneurons of protractor muscles of the tongue. For visualizing BDA, sections were treated with avidin–biotin complex and a nickel-enhanced DAB chromogen reaction. The position of labeled neurons was reconstructed with a Neurolucida equipment. Morphologically heterogeneous populations of neurons were detected bilaterally, the majority of them were distributed ipsilateral to the site of injection and extended 1200 μm in rostral and 500 μm in caudal directions. Labeled neurons were found in the rhombencephalic reticular formation, the vestibular nuclei, the nucleus prepositus hypoglossi, the nucleus of solitary tract, the spinal nucleus of trigeminal nerve and the dorsal column nuclei. Our results indicate that the majority of last-order premotor interneurons related to protractor muscles of the tongue are located in the reticular formation of the brainstem. Since this area also receives a significant input from the vestibular system and from proprioceptive fibers, the last-order premotor interneurons presented here may be the target of convergence of sensory modalities involved in prey-catching behavior.

The aim of the present study was to examine the impact of moderate and profound hyperventilation on regional cerebral blood flow (rCBF), oxygenation and metabolism.Twelve anesthetized pigs were subjected to moderate (mHV) and profound (pHV) hyperventilation (target arterial pO(2): 30 and 20 mmHg, respectively) for 30 min each, after baseline normoventilation (BL) for 1 h. Local cerebral extracellular fluid (ECF) concentrations of glucose, lactate, pyruvate and glutamate as well as brain tissue oxygenation (p(ti)O(2)) were monitored using microdialysis and a Licox oxygen sensor, respectively. In nine pigs, regional cerebral blood flow (rCBF) was also continuously measured via a thermal diffusion system.Both moderate and profound hyperventilation resulted in a significant decrease in rCBF (BL: 37.9+/-4.3 ml/100 g/min; mHV: 29.4+/-3.6 ml/100 g/min; pHV: 23.6+/-4.7 ml/100 g/min; p0.05) and p(ti)O(2) (BL: 22.7+/-4.1 mmHg; mHV: 18.9+/-4.9 mmHg; pHV: 13.0+/-2.2 mmHg; p0.05). A p(ti)O(2) decrease below the critical threshold of 10 mmHg was induced in three animals by moderate hyperventilation and in five animals by profound hyperventilation. Furthermore, significant increases in lactate (BL: 1.06+/-0.18 mmol/l; mHV: 1.36+/-0.20 mmol/l; pHV: 1.67+/-0.17 mmol/l; p0.005), pyruvate (BL: 46.4+/-7.8 micromol/l; mHV: 58.0+/-10.3 micromol/l; pHV: 66.1+/-12.7 micromol/l; p0.05), and lactate/glucose ratio were observed during hyperventilation. (Data are presented as mean+/-S.E.M.)Both moderate and profound hyperventilation may result in insufficient regional oxygen supply and anaerobic metabolism, even in the uninjured brain. Therefore, the use of hyperventilation cannot be considered as a safe procedure and should either be avoided or used with extreme caution.

Vestibular nerve branches innervating the sensory epithelia of the three semicircular canals or of the three otolith organs of frogs were selectively labeled in-vitro with biocytin. Labeled afferent fibers from the semicircular canals, utricle, and lagena were encountered in each of the four vestibular nuclei and their projections overlapped considerably. Saccular afferent fibers projected to the dorsal (acoustic) nuclei and smaller projections to the vestibular nuclei were regionally restricted. Per semicircular canal or otolith organ about equal numbers (11-14) of medium sized vestibular neurons (between 7.5 and 17 microm in diameter) were dye-coupled to afferent fibers. Most of these dye-coupled vestibular neurons were located in the lateral and descending vestibular nuclei between the VIIIth and IXth nerves. The superior vestibular nucleus was relatively free of dye-coupled vestibular neurons. The location of this subpopulation of central vestibular neurons supports the notion that these neurons are part of a particular vestibulospinal pathway. In addition, from each of the canal and/or otolith organs about 3-4 efferent vestibular neurons were labeled retrogradely. These neurons (between 15 and 26 microm in diameter) were located ventral to the vestibular nuclear complex. The branching of efferent vestibular neurons was shown by the presence of neurons that were double labeled by two different fluorescent dyes applied in the same experiment to the anterior and posterior ramus of the same VIIIth nerve, respectively. The branching of these efferent neuron axons explained the presence of collaterals and terminals in the sensory epithelia of a number of untreated ipsilateral endorgans.

The function of the neuronal isoform of nitric oxide synthase (nNOS) was studied by comparing the effects of the specific nNOS blocker 7-nitro indazole monosodium salt (7-NINA) with that of the general NOS inhibitor N G -nitro- l -arginine (L-NA) in isolated rat basilar arteries (BAs). 7-NINA had no significant effect on the resting tone of the vessels, while both L-NA and 1 H -[1,2,4]oxadiazolo[4,3- a ]quinoxalin-1-one (ODQ), a selective inhibitor of the soluble guanylyl cyclase, induced contraction. The relaxant effect of bradykinin was attenuated in the presence of L-NA but was not changed by 7-NINA. In contrast, 7-NINA markedly reduced the acetylcholine-induced, endothelium-dependent relaxation. These results demonstrate that nNOS contributes significantly to the relaxant effect of acetylcholine, indicating the functional importance of this isoenzyme in the cerebrovascular endothelium.

Investigating the cellular events in the pituitary gland, the intracellular cyclic AMP (cAMP) of the neural lobe (NL), intermediate lobe (IL), the inner (IZ-AL) and outer zone (OZ-AL) of the anterior lobe (AL) have been measured during the suckling stimulus. Ten-minutes suckling, parallel to the elevation of plasma PRL, induced a significant increase of cAMP concentration in the IZ-AL. In contrast, 10- and 30-min suckling resulted in a decrease of cAMP level in the NL. Changes in cAMP of the OZ-AL and the IL as well as in the plasma level of alpha-MSH could not be detected. These region-specific changes of cAMP in the pituitary gland during suckling stimulus seems to be related to interacting neuroendocrine signals delivered concomitantly from the hypothalamus and from the NIL to the IZ-AL.

Anxiety is a key symptom of the cocaine withdrawal syndrome in human addicts, and it is considered to be one of the major factors in precipitating relapse to chronic cocaine abuse. Corticotropin-releasing factor (CRF) plays an important role in the pathophysiology of anxiety and depression, and it may also be involved in the acute behavioral and neuroendocrine actions of cocaine. The role of endogenous CRF in cocaine withdrawal-induced anxiety was investigated in the present study. Animals were subjected to chronic cocaine (20 mg/kg, intraperitoneally, once a day for 14 days) administration. Rats tested 30 min after the last cocaine injection did not show withdrawal anxiety on the elevated plus maze or any alterations in brain CRF levels. Withdrawal (48 h) from chronic cocaine administration produced an intense anxiety-like behavior characterized by decreased open arm exploration. Immunoreactive CRF (CRF-LI) levels were selectively altered in the hypothalamus, in the amygdala and in the basal forebrain structures at the time of the behavioral anxiety, reflecting an increased activity of brain CRF systems. Daily intracerebroventricular (i.c.v.) pretreatment with an immunoserum raised against CRF completely prevented the development of anxiety induced by cocaine withdrawal. These data suggest that extrahypothalamic-limbic CRF hypersecretion may be involved in the development of anxiety related to cocaine withdrawal and that the CRF system may be a useful target for new pharmacotherapies for cocaine withdrawal and relapse.

The aim of this work was to study whether the vestibular afferent fibers establish direct connections with the motoneurons of glossopharyngeal and vagus nerves of the frog, Rana esculenta. In anaesthetized animals the vestibulocochlear nerve and the common root of glossopharyngeal-vagus and accessory (IX-X-XI) nerves were simultaneously labeled with fluorescein dextran amine (vestibulocochlear nerve) and tetramethylrhodamine dextran amine (IX-X-XI). With a confocal laser scanning microscope we could detect close appositions between the vestibular afferent fibers and somatodendritic components of the general and special visceral motoneurons of the ambiguus nucleus of IX-X nerves. The direct impulse transmission may provide a quick and immediate response of cardiovascular and gastrointestinal system upon body displacement.

We have previously demonstrated that resiniferatoxin functions in adult rats as an ultrapotent analog of capsaicin. In adults, capsaicin excites and then desensitizes a specific population of sensory neurons; when administered to neonates capsaicin causes degeneration of these neurons. We report here that treatment of newborn rats with resiniferatoxin caused a substantial (47%) loss of dorsal root ganglia neurons in adults and an almost complete loss of calcitonin gene related peptide-like immunoreactivity in both dorsal root ganglia and gasserian ganglia. The animals were unresponsive to noxious chemical stimuli and showed marked diminution (88%) of their neurogenic inflammatory response. Resiniferatoxin was at least 2 orders of magnitude more potent than capsaicin for inducing neurodegeneration in the neonates. Specific resiniferatoxin binding, thought to represent capsaicin receptors, decreased 80-90% in membranes from dorsal root ganglia and 50-70% in membranes from gasserian ganglia of adult rats treated neonatally with resiniferatoxin. The affinity for the residual binding decreased. We speculate that subpopulations of sensory neurons differ in susceptibility to neonatal resiniferatoxin treatment. Resiniferatoxin promises to be a useful probe to explore mechanisms of sensorotoxin-induced degeneration for subpopulations of capsaicin-sensitive sensory neurons.

The circuits that generate rhythmic locomotor activities are located in the ventromedial area of the lumbar spinal cord and comprise commissural interneurons necessary for left-right alternation during walking movements. In this study we injected biotinylated dextran amine (BDA) into the ventromedial gray matter of the lumbar spinal cord of neonatal rats to label commissural interneurons. Anterogradely labeled axons arose from the site of injection, crossed the midline in the anterior commissure and arborized extensively in the contralateral ventral horn of the spinal cord. The presence of neurotransmitter systems in labeled axon terminals of commissural interneurons was investigated by using antibodies raised against specific transmitter-related proteins. Boutons potentially containing inhibitory amino acids were identified by applying glutamic acid decarboxylase (GAD65/67) and glycine transporter 2 antibodies. Out of 1146 BDA-labeled axon terminals, 663 boutons were assumed on this basis to be inhibitory; 76% of these terminals were immunoreactive for glycine transporter, 53% were immunoreactive for GAD and about 30% of inhibitory boutons might contain both inhibitory amino acids. Boutons potentially containing putative excitatory neurotransmitter were revealed with antibodies raised against vesicular glutamate transporters 1 and 2. Out of 590 BDA-labeled boutons about one fourth (158) were immunoreactive for glutamate transporters. These mammalian commissural interneurons are compared to the glycinergic commissural interneurons in the swimming CPGs of lamprey and the Xenopus tadpole. Our results show that commissural interneurons in the mammalian spinal cord form a heterogeneous group including glutamatergic excitatory and GABAergic and glycinergic inhibitory neurons.

The ventrolateral cell column of the midbrain periaqueductal gray matter (vl-PAG) plays a major role in the attenuation of pain behaviour. It is established that this effect is exerted via modulation of neuronal activities in the rostral ventromedial medulla (RVM). Until recently it has been generally accepted that the vl-PAG exerts its modulatory effects upon RVM neurons through a direct monosynaptic pathway. However, recent data suggest that an additional indirect, di- or polysynaptic pathway may also exist. Using in vivo intracellular recordings we tested this hypothesis, by studying synaptic responses of somatosensory receptive RVM neurons evoked by electric stimulation of the vl-PAG in rats. RVM neurons were regarded as somatosensory receptive if they responded to electrical stimulation of the sciatic nerve. Most of the recorded RVM cells were excited by vl-PAG stimulation. Some of them responded with a short onset latency (3.6+/-0.9 ms) corresponding to monosynaptic excitation. All of these neurons were also excited by sciatic nerve stimulation at nociceptive intensities. In contrast to this, another proportion of the recorded RVM neurons responded with a four times longer (14.8+/-3 ms) onset latency to the vl-PAG stimulation, corresponding to polysynaptic modulation. All of these neurons were inhibited by sciatic nerve stimulation. The findings show that RVM neurons receive heterogeneous monosynaptic and polysynaptic inputs from the vl-PAG. The results also suggest that the monosynaptic and polysynaptic pathways modulate the activity of functionally distinct groups of RVM neurons.

Adequate treatment of cancer pain remains a significant clinical problem. To reduce side effects of treatment, intrathecal and epidural routes of administration have been used where appropriate to reduce the total dose of agent administered while achieving regional control. Resiniferatoxin (RTX), an ultrapotent capsaicin analog, gives long-term desensitization of nociception via C-fiber sensory neurons. We evaluate here the analgesic effect on rats of epidurally administered RTX, using latency of response to a thermal stimulus in unrestrained animals. Results were compared with those for systemically administered RTX. Vehicle or graded doses of RTX were injected subcutaneously (s.c.) or through an indwelling lumbar (L4) epidural catheter as a single dose. Both routes of application of RTX produced profound thermal analgesia, reaching a plateau within 4–6 h and showing no restoration of pain sensitivity over 7 days. Vehicle was without effect. For the epidural route, the effect was selective as expected for the targeted spinal cord region, whereas the subcutaneous administration of RTX had a generalized analgesic effect. At doses yielding a tripling of back paw withdrawal latency, epidural treatment was 25-fold more effective than the subcutaneous route of application. Consistent with the regional selectivity of the lumbar epidural route, the front paws showed no more effect than by systemic RTX treatment. Binding experiments with [ 3 H ]RTX provided further evidence of the segmental desensitization induced by epidural RTX. We conclude that epidural administration of RTX at the lumbar spinal level produces profound, long-lasting, segmental analgesia to C-fiber mediated pain in the rat.

Substance P immunostaining was quantified on sections from the 4th-5th lumbar and midthoracic spinal segments of rats at the peak of hyperalgesia following ultraviolet irradiation-induced inflammation of one hindpaw. The area of the immunostaining in the lumbar dorsal horn was significantly decreased on both sides by 50%, while in the thoracic spinal cord, it was increased by 18% on the contralateral and stayed unchanged on the ipsilateral side.

Vanilloid (capsaicin) receptors were visualized by [3H]resiniferatoxin (RTX) autoradiography in the brain of newborn as well as adult (both control and colchicine-treated) rats. Specific labelling was seen in the brain stem only, in the nucleus of the solitary tract extending into the area postrema and the spinal sensory nucleus of the trigeminal nerve. Also, a strong signal was seen in the dorsal horn, dorsal root, trigeminal and nodose ganglia. Membranes obtained from the cervical, thoracic, and lumbar segments of the spinal cord showed similar affinities for RTX and likewise for capsaicin and capsazepine; maximal receptor density was similar in the cervical and thoracic segments (approximately 70 fmol/mg protein) but was twice as high in the lumbar segment. 24 h after ligation of the vagal or the sciatic nerves, a strong accumulation of specific RTX binding sites was observed mainly proximal to the ligature, implying intraaxonal receptor transport from the nodose and dorsal root ganglia, respectively, to the periphery. Systemic (s.c.) vanilloid treatment depleted specific [3H]RTX binding sites from the brain stem, the sensory (dorsal root as well as trigeminal) ganglia, and the spinal cord. RTX was approximately 200-fold more potent than capsaicin for eliminating vanilloid receptors from the spinal cord. The present results suggest a discrete expression of vanilloid receptors in the brain stem (sensory nuclei); although intrinsic vanilloid receptor-expressing neurons are thought to exist in the rat brain, they remain undetected by the present [3H]RTX autoradiography methodology.

Autoradiographic mapping using [3H]resiniferatoxin (RTX) revealed high densities of vanilloid binding sites over areas (Rexed laminae I and II) in pig spinal cord known to be rich in central terminals of capsaicin-sensitive neurons. Also, high affinity [3H]RTX binding was detected in membranes obtained from the corresponding areas: apparent binding affinity and cooperativity but not the maximal receptor density was influenced by the assay conditions (temperature, buffer composition). No specific binding could be detected in other areas of the spinal cord by either methodology suggesting that the vanilloid receptors are present exclusively on central terminals of the capsaicin-sensitive neurons. We conclude that [3H]RTX autoradiography may afford a novel neurochemical approach to detect localized changes in vanilloid receptor expression.

Corticotropin-releasing factor (CRF) may mediate some of the neuroendocrine and behavioral responses to cocaine. In this study, the distribution of CRF-like immunoreactivity (CRF-LI) was determined in the hypothalamus and in several extrahypothalamic brain regions after acute cocaine administration in handled rats. CRF-LI decreased dose-dependently with cocaine administration in the hypothalamus and in the basal-forebrain structures. A small dose of cocaine (7.5 mg/kg) decreased CRF-LI in the hippocampus and in the frontal cortex. A significant, selective, dose-dependent increase in CRF-LI was found in the amygdala after cocaine injection. None of the investigated doses of cocaine altered CRF-LI in the striatum. These results suggest that acute cocaine administration alters brain CRF systems to contribute behavioral and neuroendocrine responses to cocaine.

Immunoreactive oxytocin (IR-OXT) detected in extracts of human lumbar sympathetic paravertebral ganglia was characterized by high-performance liquid chromatography (HPLC). The immunoreactive substance was found to elute at the same position as the reference preparation of oxytocin (OXT). The results revealed the presence of chromatographically identified OXT in human sympathetic ganglia.

Immunoreactive oxytocin levels were measured in the mouse hippocampal tissue (h-OXT). Acute morphine treatment increased h-OXT, which effect was reversed by naloxone. In mice rendered tolerant to and dependent on morphine h-OXT was lower than in placebo pellet-implanted control mice. In the tolerant/dependent animals naloxone resulted in precipitated withdrawal syndrome which was associated with a slight increase in h-OXT. The data indicate that h-OXT is affected by acute morphine treatment and by morphine tolerance/dependence and raises the possibility that h-OXT participates in the adaptive response of the organism towards narcotic drugs.

Oxytoxin-like immunoreactivity (IR-OXT) was detected in extracts of human spinal L5 and Gasserian ganglia by a radioimmunoassay (RIA) specific to oxytocin (OXT) and was identified by high-performance liquid chromatography (HPLC). One of the two immunoreactive peaks obtained on HPLC was found to elute at the same position as the OXT standard. The results reveal the presence of chromatographically identified OXT immunoreactivity in human sensory ganglia.