Search

Alpha-chloralose (AC) is used as a rodenticide as well as an anesthetic agent in laboratory animals. It was previously also used as an avicide. Detection of AC in blood samples or in body tissues collected postmortem is key for the diagnosis of clinical cases and a requirement for surveillance of secondary toxicosis, including potential cases in wild animals. Reports on poisoning of humans and non-laboratory animals confirmed by the detection of AC or its metabolites are available, however poisoning of domestic animals are rarely available. Furthermore, reports on clinical cases in domestic animals rarely report quantifications of AC in blood or body tissues. The present study describes the validation of a quantitative ultra high performance liquid chromatography--tandem mass spectrometry (UHPLC--MS-MS) method that can be used in cases of suspected AC poisoning in cats. The validation study showed the method to be fit for purpose. In serum, the limit of quantification was 100 ng/mL and the limit of detection was 30 ng/mL. The new analytical method was applied on blood samples collected from 20 individual cats with a preliminary clinical diagnosis of acute AC poisoning. AC was confirmed in all 20 feline blood samples, and the concentration range of AC was 538–17,500 ng/mL. The quantitative method developed in this study was found to be a fast and selective method for confirmation of AC poisoning using blood samples from cats.

Spontaneous neural activity has been widely adopted to construct functional connectivity (FC) amongst distant brain regions. Although informative, the functional role and signaling mechanism of the resting state FC are not intuitive as those in stimulus/task-evoked activity. In order to bridge the gap, we investigated anesthetic modulation of both resting-state and sensory-evoked activities. We used two well-studied GABAergic anesthetics of varying dose (isoflurane: 0.5–2.0% and α-chloralose: 30 and 60 mg/kg∙h) and recorded changes in electrophysiology using a pair of laminar electrode arrays that encompass the entire depth of the bilateral somatosensory cortices (S1fl) in rats. Specifically, the study focused to describe how varying anesthesia conditions affect the resting state activities and resultant FC between bilateral hemispheres in comparison to those obtained by evoked responses. As results, isoflurane decreased the amplitude of evoked responses in a dose-dependent manner mostly due to the habituation of repetitive responses. However, α-chloralose rather intensified the amplitude without exhibiting habituation. No such diverging trend was observed for the spontaneous activity, in which both anesthetics increased the signal power. For α-chloralose, overall FC was similar to that obtained with the lowest dose of isoflurane at 0.5% while higher doses of isoflurane displayed increased FC. Interestingly, only α-chloralose elicited relatively much greater increases in the ipsi-stimulus evoked response (i.e., in S1fl ipsilateral to the stimulated forelimb) than those associated with the contra-stimulus response, suggesting enhanced neuronal excitability. Taken together, the findings demonstrate modulation of the FC profiles by anesthesia is highly non-linear, possibly with a distinct underlying mechanism that affects either resting state or evoked activities differently. Further, the current study warrants thorough investigation of the basal neuronal states prior to the interpretation of resting state FC and evoked activities for accurate understanding of neural signal processing and circuitry.

The aim of this study was to assess the effects of radiofrequency electromagnetic field (RF EMF) on heart rate variability (HRV) in rabbits with intensity slightly exceeding the limits for occupations. Totally 21 New Zealand white rabbits divided into two groups were used in this double-blind study. The first group of animals without general anesthesia was subjected to HRV examination under exposure to a device generated RF EMF source (frequency 1788 MHz, intensity 160 V/m, lasting 150 min.). The second group (premedications + α chloralose mg/kg) underwent the same protocol under the exposure to the real RF EMF signal from the base stations of mobile providers (frequency range 1805 – 1870 MHz - corresponding to the downlink signal of Slovak mobile providers, 160 V/m, 150 min., respectively). Individual 5 min records were used to analyze the HRV parameters: heart rate and root Mean Square of the Successive Differences (rMSSD) for time domain analysis and spectral powers in the low (LF-VFS) and high frequency (HF-VFS) bands for frequency domain analysis. Our study revealed the increased in HRV parameters (HF-HRV, rMSSD) associated with lower heart rate indicating increased cardiac vagal control under the exposure to RF EMF in experimental methods.

Objectives The aim of this study was to describe the clinical picture in cats with alpha-chloralose (AC) intoxication and to confirm AC in serum from suspected cases of AC poisoning. Methods Suspected cases of AC poisoning were identified in patient records from a small animal university hospital from January 2014 to February 2020. Clinical signs of intoxication described in respective records were compiled, the cats were graded into four intoxication severity scores and hospitalisation time and mortality were recorded. Surplus serum from select cases in late 2019 and early 2020 was analysed to detect AC with a quantitative ultra-high performance liquid chromatography tandem mass spectrometry analysis, and the AC concentration was compared with the respective cat’s intoxication severity score. Results Serum from 25 cats was available for analysis and AC poisoning was confirmed in all. Additionally, 78 cats with a clinical suspicion of AC intoxication were identified in the patient records, most of which presented from September to April. The most common signs of intoxication were ataxia, tremors, cranial nerve deficits and hyperaesthesia. The prevalence of clinical signs and intoxication severity differed from what has previously been reported, with our population presenting with less severe signs and no deaths due to intoxication. The majority had a hospitalisation time Conclusions and relevance This study describes the clinical signs and prognosis in feline AC intoxication. There were no mortalities in confirmed cases, indicating that AC-poisoned cats have an excellent prognosis when treated in a timely manner. Recognition of AC intoxication as a differential diagnosis for acute onset of the described neurological signs in areas where AC exposure is possible may influence clinical decision-making and help avoid excessive diagnostic procedures. A severe clinical picture upon presentation could be misinterpreted as a grave prognosis and awareness about AC poisoning may avoid unnecessary euthanasia.

Purpose: identification of pathological changes in the liver at early stages of the effect of clozapine and clozapine combined with ethanol to improve the diagnosis of this acute exposure.Materials and methods. The experiments were carried out on outbred male rats (n=15). The animals were split into 3 groups, 5 animals in each: 1) control (intact rats); 2) clozapine, 3) clozapine and ethanol. Clozapine at a dose of 150 mg/kg of the animal’s body weight was administered orally under anesthesia (chloralose), clozapine (150 mg/kg) with ethanol at a dose of 5 ml/kg — orally. Rats were removed from the experiment after 3 hours by decapitation. Samples of liver tissue were studied using light microscopy. During histological examination, the following morphological signs were evaluated: blood circulation disorder (presence of venous, capillary, and arterial congestion, appearance of venous hemorrhage), absence of nuclei staining, cytoplasm lumpiness, cytoplasm vacuolation, presence of necrosis foci, appearance of cellular response (a large quantity of leukocytes per power field). The assessment was done using the F-test. Forensic chemical analysis was carried out using a high-performance liquid chromatograph equipped with a mass detector. The content of clozapine, norclozapine, and clozapine-N-oxide in blood serum and liver homogenate of rats was evaluated.Results. In the control group, no pathological changes in liver tissue were observed. 3 hours after clozapine administration, venous congestion was detected. 3 hours after administration of clozapine and ethanol combination, venous congestion and absence hepatocyte nuclei staining were observed. It was found that after combined administration of clozapine and ethanol, the blood serum clozapine was 1.94-fold and clozapine-N-oxide was 2.3-fold lower than when clozapine was administered as a single drug while the content of norclozapine showed no difference.Conclusion. Changes detected in the liver, together with changes in other organs and findings of the forensic chemical analysis, allow establishing the fact of clozapine poisoning. Understanding of the processes going on in the body during such intoxications helps rendering medical assistance to victims more effectively.

Purpose Sensitivity and specificity of blood oxygenation level–dependent (BOLD) functional MRI (fMRI) is sensitive to magnetic field strength and acquisition methods. We have investigated gradient‐echo (GE)‐ and spin‐echo (SE)‐BOLD fMRI at ultrahigh fields of 9.4 and 15.2 Tesla. Methods BOLD fMRI experiments responding to forepaw stimulation were performed with 3 echo times (TE) at each echo type and B 0 in α‐chloralose–anesthetized rats. The contralateral forelimb somatosensory region was selected for quantitative analyses. Results At 9.4 T and 15.2 T, average baseline T 2 * (n = 9) was 26.6 and 17.1 msec, whereas baseline T 2 value (n = 9) was 35.7 and 24.5 msec, respectively. Averaged stimulation‐induced ΔR 2 * was –1.72 s–1 at 9.4 T and –3.09 s–1 at 15.2 T, whereas ΔR 2 was –1.19 s–1 at 9.4 T and –1.97 s–1 at 15.2 T. At the optimal TE of tissue T 2 * or T 2, BOLD percent changes were slightly higher at 15.2 T than at 9.4 T (GE: 7.4% versus 6.4% and SE: 5.7% versus 5.4%). The ΔR 2 * and ΔR 2 ratio of 15.2 T to 9.4 T was 1.8 and 1.66, respectively. The ratio of the macrovessel‐containing superficial to microvessel‐dominant parenchymal BOLD signal was 1.73 to 1.76 for GE‐BOLD versus 1.13 to 1.19 for SE‐BOLD, indicating that the SE‐BOLD contrast is less sensitive to macrovessels than GE‐BOLD. Conclusion SE‐BOLD fMRI improves spatial specificity to microvessels compared to GE‐BOLD at both fields. BOLD sensitivity is similar at the both fields and can be improved at ultrahigh fields only for thermal‐noise–dominant ultrahigh‐resolution fMRI.

WOS: 000403692800006, Four new chiral Schiff base ligands (3a-4a, 3b-4b) have been prepared using aminoisopropylidene derivatives of glucose and aminochloralose derivatives of mannose. The synthesized compounds 1 and 2 were characterized by nuclear magnetic resonance (H-1 and C-13 NMR), Fourier transform infrared spectroscopy (FTIR), UV/visible spectroscopy, and elemental analysis. A total of eight Schiff base compounds (3a-6a, 3b-6b) were tested for antimicrobial activity against Escherichia coli ATCC 12228, Staphylococcus aureus ATCC 6538, Klebsiella pneumoniae CCM 2318, Pseudomonas aeruginosa ATCC 27853, Salmonella typhimurium CCM 5445, Enterococcus faecalis ATCC 29212, Bacillus subtilis ATCC 6633, and Candida albicans ATCC 10239 and exhibited a range of activities.

In this study, D-glucose and D-galactose were used as starting sugar. Firstly, the experimental works were constituted on the simple synthesis of the 6-amino derivatives of 1,2-O-(R)-trichloroethylidene-α-D-glucofuranose (α-chloralose) and 1,2-O-(S)-trichloroethylidene-α-D-galactofuranose. The trichloroethylidene acetals of D-glucose and D-galactose were prepared from the reaction of sugars and anhydrous chloral. Tosyl derivates of trichloroethylidene acetals of D-glucose and D-galactose were obtained through the reaction of these compounds with p-toluenesulfonylchloride and then, these tosyl derivates were converted to the azide form by following nuclephilic substitution reaction. Obtained azide derivates were reacted with triphenylphosphine as reducing agent in order to get the related amino-sugars derivates. Also the same amino-sugars were synthesized in the C-3 hydroxy groups are methylated to derivatives by the same method. Then, in order to synthesize Schiff bases; the homogeneous mixture aqua form of aldehyde (salicylaldehyde or 3,5-ditbutylsalicylaldehyde) in MeOH was added dropwise into the solution of amino sugar derivative in 5mL of MeOH. Schiff bases were synthesized at the last of reaction. For to perform Henry reaction; to a solution of shiff bases as a ligand and solvent at the given temperature was added Cu(OAc)2.nH2O. Then the aldehyde(4-nitrobenzaldehit) and nitromethane were added into the solution. Subsequential of the completion, to afford the desired Henry product the solvent was evaporated and the residue was purified by column chromatography using hexane:ethyl acetate. HPLC using a Chiralcel OD-H column used to determine the enantiomeric excess values. Spectroscopic analysis, like 1H-NMR, IR and13C-NMR, helped us to learn the structures of all the compounds., Bu çalışmada D-glukoz ve D-galaktoz başlangıç şekerleri olarak kullanılmıştır. Öncelikle deneysel çalışmalar, 1,2-O-(R)-trikloroetiliden-α-D-glukofuranoz (α-kloraloz) ve 1,2-O-(S)-trikloroetiliden-α-D-galaktofuranoz'un 6-amino türevlerinin sentezleri üzerinde oluşturulmuştur. D-glukoz ve D-galaktoz'un trikloroetiliden acetalleri, susuz kloral ve şekerlerle reaksiyonu sonucu hazırlanmıştır. D-glukoz ve D-galaktoz'un trikloroetiliden acetallerinin tosil türevleri p-toluensülfonilklorür ile reaksiyon sonucu elde edimiştir ve daha sonra bu tosil türevleri nükleofilik yerdeğiştirme tepkimesi ile azit formuna dönüştürülmüştür. Elde edilen azit bileşikleri, ilgili amino şeker türevini elde etmek için indirgeyici reaktif olan trifenilfosfin ile reaksiyona sokulmuştur. Ayrıca bu amino şekerlerin C-3 hidroksil grupları metillenmiş olan türevleride benzer yöntemle sentezlenmiştir. Daha sonra Schiff bazı sentezlemek amacı ile, MeOH içinde aldehit (salisilaldehit veya 3,5-ditbutilsalisilaldehit) çözeltisi 5 ml MeOH'deki amino şeker türevi çözeltisi içine damlatılarak ilave edilmiştir. Henry reaksiyonunu gerçekleştirmek amacıyla belirtilen sıcaklıkta ligant olan Schiff bazı ve çözgen, Cu(OAc)2.nH2O üzerine eklenmiştir. Daha sonra aldehit (4-nitrobenzaldehit) ve nitrometan bu çözeltiye ilave edilmiştir. Reaksiyon tamamlandıktan sonra solvent buharlaştırılarak istenen Henry ürünü, hekzan ve etilasetat kullanılarak kolon kromatografisi ile saflaştırılmıştır. Enantiyomerik fazlalık değerleri Chiralcel OD-H kolon kullanılarak HPLC'de belirlenmiştir. Bütün bileşiklerin yapıları 1H-NMR, 13C-NMR ve IR gibi spektroskopik yöntemlerle belirlenmiştir.

In this research, D-mannose, D-galactose and D-glucose were used as starting sugars. Starting from these monosaccharides, the synthesis and characterization of acrylic and methacrylic polymers were investigated. As a result of anhydrous chloral and D-galactose reaction, 1,2-O-trichloroethylidene-α-D-galactofuranose was obtained. 5,6-O-isopropylidene-1,2-O-trichloroethylidene-α-D-galactofuranose was obtained, 2-O- trichloroethylidene -α-D-galactofuranose and 2,2-dimethoxypropane. In the next stage, 5,6-O-isopropylidene-1,2-O-trichloroethylidene-α-D-galactofuranose and epichlorohydrine were reacted and the result of this reaction epoxy ring was added to number 3 carbon. Epoxy rings may be opened with the base catalyst. The use of this property was performed ring opening reactions with acrylic and methacrylic acid. In this way, sugar-based monomers were obtained and investigated. Likewise D-glucose and D-mannose were used as starting sugar.5,6-O-isopropylidene-1,2-O-trichloroethylidene-α-D-glucofuranose and 5,6-O-isopropylidene-1,2-O-trichloroethylidene-α-D-mannofuranose were obtained. The steps of D-galactose reaction were repeated for other sugars., Bu araştırmada D-mannoz, D-galaktoz ve D-glukoz başlangıç şekerleri olarak kullanıldı. Bu monosakkaritlerden yola çıkılarak yeni akrilik ve metakrilik polimerlerin sentezi ve karakterizasyonu üzerine çalışılmıştır. D-galaktoz'un susuz kloralle tepkimesi sonucu 1,2-O-trikloroetiliden-α-D-galaktofuranoz elde edildi. 1,2-O-trikloroetiliden-α-D-galaktofuranoz ile 2,2-dimetoksipropan tepkimeye sokularak 5,6-O-izopropiliden-1,2-O-trikloroetiliden-α-D-galaktofuranoz oluşturuldu. Bir sonraki basamakta 5,6-O-izopropiliden-1,2-O-trikloroetiliden-α-D-galaktofuranoz'un epiklorohidrin ile tepkimesi sonucu 3 numaralı karbona epoksi halkası bağlandı. Epoksi halkasının bazik ortamdaki açılma tepkimesinin özelliğinden faydalanarak akrilik ve metakrilik asitler ile halka açılma tepkimesi gerçekleştirildi. Benzer şekilde D-glukoz ve D-mannoz' dan başlayarak sırasıyla 5,6-O-izopropiliden-1,2-O-trikloroetiliden-α-D-glukofuranoz ve 5,6-O-izopropiliden-1,2-O-trikloroetiliden-α-D-mannofuranoz oluşturuldu. D-galaktoz'un reaksiyon basamakları bu şekerler için de aynen tekrarlandı.

Objective: to detect lung morphological changes in acute intoxications with clozapine, ethanol, and their combination 3 and 24 hours after poisoning. Materials and methods. Experiments were carried out in outbred male rats weighing 270—300 g. Clozapine was given in a dose of 250 mg per kg animal body weight under chloralose anesthesia. Following 3 and 24 hours, the animals were withdrawn from the experiment by decapitation. Lung histological sections from 6 rats that had received oral clozapine 250 mg/kg, 6 rats that had oral ethanol 8.6 ml/kg, and 6 rats that had a combination of ethanol and clozapine orally in the above doses were examined 3 hours after intoxication. Those from 18 rats that had been orally given the similar agents in the above doses and withdrawn from the experiment were also investigated 24 hours after drug administration. The sections were compared with those from 6 rats that had not received the above agents. Nonparametric methods (χ2 test) were used for statistical processing. The investigators assessed the following morphological signs: circulatory disorders (plethora, hemorrhages), interstitial and alveolar edema, damage to the bronchial and alveolar epithelium and to the endothelium, and a cell reaction. The differences were considered significant at p

Resting-state functional magnetic resonance imaging (rsfMRI) is a translational imaging method with great potential in several neurobiologic applications. Most preclinical rsfMRI studies are performed in anesthetized animals, but the confounding effects of anesthesia on the measured functional connectivity (FC) are poorly understood. Therefore, we measured FC under six commonly used anesthesia protocols and compared the findings with data obtained from awake rats. The results demonstrated that each anesthesia protocol uniquely modulated FC. Connectivity patterns obtained under propofol and urethane anesthesia were most similar to that observed in awake rats. FC patterns in the α-chloralose and isoflurane-medetomidine combination groups had moderate to good correspondence with that in the awake group. The FC patterns in the isoflurane and medetomidine groups differed most from that in the awake rats. These results can be directly exploited in rsfMRI study designs to improve the data quality, comparability, and interpretation, published version, peerReviewed

It has previously been proved that sodium nitrite, infused prior to coronary artery occlusion or before reperfusion, results in marked antiarrhythmic effect in anaesthetized dogs. We have now examined whether this protection involves the modulation of gap junction (GJ) function by nitric oxide (NO), derived from nitrite administration under ischaemic conditions. Two groups of chloralose and urethane anaesthetized dogs, each containing 13 animals, were subjected to a 25 min period occlusion of the left anterior descending (LAD) coronary artery, followed by reperfusion. One group was infused with sodium nitrite (0.2 μmol/kg/min, i.v.), the other group with saline 10 min prior to reperfusion. The severities of arrhythmias and of ischaemia (epicardial ST-segment, total activation time), parallel with changes in myocardial tissue impedance, a measure of electrical coupling of gap junctions, were assessed during the experiments. Compared to the controls, nitrite infusion administered prior to reperfusion significantly attenuated the severity of ischaemia, the ischaemia-induced impedance changes and, consequently, the severity of arrhythmias, occurring during the 1B phase of the occlusion, and increase survival following reperfusion (0% vs. 85%). It is concluded that the marked antiarrhythmic effect of sodium nitrite is partly due, to the preservation of the electrical coupling of GJs by NO.

A five-year-old domestic cat with acute unexplainable neurological signs was presented for postmortem examination. Clinically, the cat showed an acute onset of ataxia, depressed mentation and continuous twitching/ seizure activity in the morning after having been outside overnight. Despite immediate treatment, the cat# progressed within 24 hours to a comatose state, opisthotonus and severe miosis unresponsive to light. Given a poor prognosis, euthanasia was elected. Gross findings were disappointing and consisted of a nonspecific lung oedema and congested lungs and spleen. Surprisingly, within the stomach and intestines, fragments of cockshafers were found. Histological examination confirmed the gross findings and additionally showed evidence of mild brain oedema, but failed to identify a cause for the severe clinical signs. In a final attempt to solve the case, a urine sample was tested for toxic substances and it was found to contain a significant amount of α-chloralose. This finding was unexpected.

Background Mechanosensitive afferents innervating the bladder increase their firing rate as the bladder fills and pressure rises. However, the relationship between afferent firing rates and intravesical pressure is not a simple linear one. Firing rate responses to pressure can differ depending on prior activity, demonstrating hysteresis in the system. Though this hysteresis has been commented on in published literature, it has not been quantified. Results Sixty-six bladder afferents recorded from sacral dorsal root ganglia in five alpha-chloralose anesthetized felines were identified based on their characteristic responses to pressure (correlation coefficient ≥ 0.2) during saline infusion (2 ml/min). For saline infusion trials, we calculated a maximum hysteresis ratio between the firing rate difference at each pressure and the overall firing rate range (or Hmax) of 0.86 ± 0.09 (mean ± standard deviation) and mean hysteresis ratio (or Hmean) of 0.52 ± 0.13 (n = 46 afferents). For isovolumetric trials in two experiments (n = 33 afferents) Hmax was 0.72 ± 0.14 and Hmean was 0.40 ± 0.14. Conclusions A comprehensive state model that integrates these hysteresis parameters to determine the bladder state may improve upon existing neuroprostheses for bladder control.

Despite positive metabolic effects, genetic deletion of protein phosphatase 1b (PTP1b) results in sympathetically mediated elevations in arterial pressure (AP) in mice. Because several PTP1b-regulated peptides also impair the baroreflex sensitivity (BRS) for control of heart rate (HR), we hypothesized that PTP1b in the solitary tract nucleus (NTS) participates in the maintenance of resting baroreflex function. To test this hypothesis, we performed acute bilateral microinjection of an allosteric PTP1b inhibitor (100 nM/120 nL) in the NTS of urethane/chloralose anesthetized Sprague-Dawley rats and assessed the BRS, responses to cardiac vagal chemosensitive fiber activation, and resting AP and HR before and after the injection. PTP1b inhibition impaired the BRS for bradycardia (n = 6; 0.93 ± 0.14 baseline vs. 0.48 ± 0.04 at 10 minutes vs. 0.49 ± 0.04 millisecond/mm Hg at 60 minutes; P0.01), with no significant effect on the BRS for tachycardia (0.30 ± 0.16 baseline vs. 0.24 ± 0.08 at 10 minutes vs. 0.24 ± 0.12 millisecond/mm Hg at 60 minutes). The reduced BRS for bradycardia was associated with a significant decrease in alpha-adrenergic responsiveness to phenylephrine at 60 minutes after PTP1b inhibition. Injection of the PTP1b inhibitor in the NTS elicited transient decreases in AP and HR in these animals. However, there was no effect of the inhibitor on depressor or bradycardic responses elicited by activation of cardiac vagal chemosensitive fibers, which converge with baroreceptor afferents in the NTS. These results suggest that PTP1b within the NTS may be a novel molecular mechanism for preservation of resting baroreflex function and provides further evidence for deleterious cardiovascular effects associated with PTP1b inhibition.

Nitric oxide (NO), a potent vasodilator and nontraditional neurotransmitter, is an important mediator of the changes in cerebral blood flow (CBF) associated with increased neuronal activity (neurovascular coupling). In the present work, we investigated the role of NO and of its newly recognized precursor, nitrite, in neurovascular coupling using a well-established rat model of somatosensory stimulation. Biological synthesis of NO of neuronal origin was inhibited pharmacologically. Following the initial uncoupling of neuronal and hemodynamic responses to somatosensory stimulation, the NO donor sodium nitroprusside, added within the range of physiological concentrations, significantly increased, but did not fully restore the functional CBF response. In contrast, nitrite at its physiological concentration fully recovered neurovascular coupling to its original magnitude. The magnitude of the effect is, however, dose-dependent. Sub-physiological concentrations of nitrite were not enough to entirely restore neurovascular coupling and supra-physiological concentrations acted more as a local vasodilator that changed resting CBF and interfered with the functional CBF response. These results suggest that nitrite can be efficiently converted into NO and utilized to support normal cerebrovascular physiology.

Our previous studies showed that electrical stimulation of the nuclei ambiguous (NA) or dorsomotor nuclei of the vagus (DMV) complex in the brain stem of spontaneously breathing pond turtles (Cyclemys fiavomarginata), anesthetized with chloralose (4 mg/100 g) and urethane (40 mg/100 g), produced a marked slowing or even cessation of the heart rate, and resulted in an immediate fall of blood pressure. Results of the present study further demonstrated that the cardioinhibitory responses could also be elicited by microinjection of monosodium glutamate (0.2-20 nl, 50 mM) into the NA/DMV complex in turtles. A two-barrel glass micropipette held in a manipulator was connected to a pneumatic pressure pump for microinjection. The glutamate-induced cardioinhibitory responses could be significantly reduced in a dose-dependent manner by pretreatment with AP-5 (a NMDA receptor antagonist, at 1-8 nmole) or CNQX (a non-NMDA receptor antagonist; at 0.1-0.8 nmole) 20 min before glutamate administration. Histochemical verification by injecting horseradish peroxidase into the cervical vagus nerves revealed that retrogradely labeled glutamatergic neurons in the NA/DMV complex were observed. These results suggest that glutamatergic receptors in the caudal medulla may mediate vagal cardioinhibitory responses in the turtle.

Background and purpose: Both ischaemia preconditioning (PC) and the intracoronary infusion of peroxynitrite (PN) suppress ischaemia and reperfusion (I/R)-induced arrhythmias and the generation of nitrotyrosine (NT, a marker of PN). However, it is still unclear whether this latter effect is due to a reduction in nitric oxide (NO) or superoxide (O2-) production. Experimental approach: Dogs anaesthetized with chloralose and urethane were infused, twice for 5 min, with either saline (control) or 100 nM PN, or subjected to similar periods of occlusion (PC), 5 min prior to a 25 min occlusion and reperfusion of the left anterior descending coronary artery. Severities of ischaemia and ventricular arrhythmias, as well as changes in the coronary sinus nitrate/nitrite (NOx) levels were assessed throughout the experiment. The production of myocardial NOx, O2- and NT was determined following reperfusion. Key results: Both PC and PN markedly suppressed the I/R-induced ventricular arrhythmias, compared to the controls, and increased NOx levels during coronary artery occlusion. Reperfusion induced almost the same increases in NOx levels in all groups, but superoxide production and, consequently, the generation of NT were significantly less in PC- and PN-treated dogs than in controls. Conclusions and implications: Since both PC and the administration of PN enhanced NOx levels during I/R, the attenuation of endogenous PN formation in these dogs is primarily due to a reduction in the amount of O2 produced. Thus, the anti-arrhythmic effect of PC and PN can almost certainly be attributed to the preservation of NO availability during myocardial ischaemia.

Autoregulation, an important feature of the cerebral circulation, is affected in many diseases. Since genetically modified mice are a fundamental tool in biomedical research, including neuro(bio)logy also in this specie measurements of cerebral autoregulation (CA) are mandatory. However, this requires anesthesia that unfortunately significantly impacts cerebral perfusion and consequently might distort CA measurements directly or by altering arterial pCO(2). The latter can be avoided by artificial ventilation but requires several control measurements of blood gases, each consuming at least 100 μl of blood or 5% of a mouse's blood volume. To avoid such diagnostic hemorrhage, we systematically analyzed the effect of different common anesthetic protocols used for rodents in spontaneously breathing mice on CA measured with Laser speckle perfusion imaging. Halothane, Isoflurane and Pentobarbital abrogated CA and Ketamin/Xylazine as well as Chloralose had a moderate reproducibility. In contrast, the rather rarely used anesthetic Ethomidate applied in low doses combined with local anesthetics had the best reproducibility. Although with this anesthesia the lower CA limit was lower than with Ketamin/Xylazine and Chloralose as reported in the handful of papers so far dealing with CA in mice, we suggest Ethomidate as the anesthetic of choice for CA measurements in spontaneously breathing mice.

The regulation of blood pressure and sympathetic outflow by the brain renin–angiotensin system in animals subjected to raised or lowered dietary Na+ intake is unclear. This study compared the mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA) responses to intracerebroventricular (i.c.v.) infusion of angiotensin II (AngII) and III (AngIII) before and after peripheral V1 receptor blockade (V1B) in α-chloralose–urethane-anaesthetized rats fed a low (0.03%, LNa+), normal (0.3%, NNa+) or high Na+ diet (3.0%, HNa+) from 4 to 11 weeks of age. The rise in MAP 2 min post AngII i.c.v. was greater in HNa+ (14 ± 3 mmHg) versus LNa+ (8 ± 1 mmHg, P < 0.05) and after AngIII i.c.v. in HNa+ (14 ± 3 mmHg) versus NNa+ (6 ± 1 mmHg, P < 0.05) and LNa+ (7 ± 1 mmHg, P < 0.05). The MAP responses to AngII and AngIII i.c.v. were abolished after V1B in LNa+, but were only attenuated in HNa+. In NNa+, V1B blunted the MAP responses to AngII and abolished those to AngIII. The MAP remained elevated 30 min after AngII in all groups, but returned to baseline levels 15 min after AngIII in NNa+ and HNa+ (P < 0.01). Twenty minutes after i.c.v. AngII, RSNA rose above baseline in HNa+ (112 ± 1%), a response not observed in the LNa+ and NNa+ groups. Twenty minutes post AngIII i.c.v., RSNA was elevated in both HNa (109 ± 2%) and NNa+ (109 ± 2%). After V1B, RSNA rose only in the HNa+ group 15 min post AngIII infusion (109 ± 1%). Together, these findings: (1) suggest that HNa+ intake augments the MAP and RSNA responses to i.c.v. AngII and AngIII; (2) highlight an important role for peripheral V1 receptors during these responses; and (3) differentiate the effects of AngII and AngIII on blood pressure and RSNA.

Most studies of the effect of cocaine on brain activity in laboratory animals are preformed under anesthesia, which could potentially affect the physiological responses to cocaine. Here we assessed the effects of two commonly used anesthetics [alpha-chloralose (alpha-CHLOR) and isofluorane (ISO)] on the effects of acute cocaine (1 mg/kg i.v.) on cerebral blood flow (CBF), cerebral blood volume (CBV), and tissue hemoglobin oxygenation (S(t)O(2)) using optical techniques and cocaine's pharmacokinetics (PK) and binding in the rat brain using (PET) and [(11)C]cocaine. We showed that acute cocaine at a dose abused by cocaine abusers decreased CBF, CBV and S(t)O(2) in rats anesthetized with ISO, whereas it increased these parameters in rats anesthetized with alpha-CHLOR. Importantly, in ISO-anesthetized animals cocaine-induced changes in CBF and S(t)O(2) were coupled, whereas for alpha-CHLOR these measures were uncoupled. Moreover, the clearance of [(11)C]cocaine from the brain was faster for ISO (peak half-clearance 15.8 +/- 2.8 min) than for alpha-CHLOR (27.5 +/- 0.6 min), and the ratio of specific to non-specific binding of [(11)C]cocaine in the brain was higher for ISO- (3.37 +/- 0.32) than for alpha-CHLOR-anesthetized rats (2.24 +/- 0.4). For both anesthetics, cocaine-induced changes in CBF followed the fast uptake of [(11)C]cocaine in the brain (peaking at approximately 2.5-4 min), but only for ISO did the duration of the CBV and S(t)O(2) changes correspond to the rate of [(11)C]cocaine's clearance from the brain. These results demonstrate that anesthetics influence cocaine's hemodynamic and metabolic changes in the brain, and its binding and PK, which highlights the need to better understand the interactions between anesthetics and pharmacological challenges in brain functional imaging studies.

The ability of intravenous cocaine to induce Fos protein expression in anesthetized rats was tested. Two anesthetic regimens commonly used for in vivo FMRI of animals, i.v. alpha-chloralose and gaseous isoflurane, were studied in separate cohorts. The first experiment included three groups that received the following treatments: saline i.v. and no anesthetic; 2 mg/kg cocaine i.v. and no anesthetic; and 2mg/kg cocaine i.v. under 36 mg/kg/h alpha-chloralose anesthesia. The second experiment had a factorial design of four groups that were either nonanesthetized or isoflurane-treated and were either given saline or cocaine (2 mg/kg, i.v.). Anesthetized rats were maintained for 2 h under 2.5-3.5% isoflurane anesthesia, while nonanesthetized rats were kept in an alternative environment for the same time period. Rats were given 2 mg/kg cocaine or saline i.v., 30 min into the test session. Rats were perfused and their brains were processed for Fos immunohistochemistry 90 min after the i.v. treatment. In both experiments, the frontal cortex and striatum of the cocaine-treated nonanesthetized rats expressed Fos in greater amounts than the saline-treated nonanesthetized rats, as expected. The alpha-chloralose treatment prevented cocaine-induced Fos expression across all eight subregions of the striatum and frontal cortex that were examined. In contrast, isoflurane only partially attenuated Fos expression in the orbital and Cg2 subregions of frontal cortex. These results suggest a strong advantage for using isoflurane, as opposed to alpha-chloralose, when studying anesthetized rats for in vivo effects of psychostimulants.

Focal ventricular tachycardia (VT) in acute myocardial ischemia is closely related to triggered activity (TA), which may be blocked by scavenging reactive oxygen species (ROS).This study analyzed effects of acutely administered ROS scavenger-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO) on VT in vivo and TA in vitro.Forty-three alpha chloralose anesthetized dogs with coronary artery occlusion were studied. Three-dimensional activation mapping helped to locate the origin of focal or reentrant VT. TEMPO (30 mg/kg intravenously) or vehicle was given. Endocardium excised from the site of origin of VT was studied using standard microelectrode techniques and measures of ROS.Reentry and focal VT induction were both highly reproducible. TEMPO blocked focal VT in 6 of 11 dogs (P.05), but 9 of 9 dogs with reentrant VT continued to have VT re-induced after TEMPO. TEMPO did not alter effective refractory period (168 +/- 3 to 171 +/- 3 ms), mean blood pressure (88 +/- 3 to 81 +/- 3 mm Hg), and size of ischemia (42% +/- 3% vs 40% +/- 4%). In vitro, TEMPO (10(-3) M, n = 14) produced no change in action potentials. Nevertheless, TA was reversibly attenuated from 5.3 +/- 1.1 to 0.4 +/- 0.4 complexes with TEMPO (n = 15, P.05). Lucigenin-enhanced chemiluminescence and dihydroethidium staining showed increased ROS in ischemic endocardium; TEMPO dramatically reduced ROS in ischemic sites.TEMPO, a scavenger of ROS, prevented triggered activity associated with focal VT during myocardial ischemia in areas of increased ROS. Antioxidant therapy may play an important role in blockade of focal VT under the conditions of myocardial ischemia.

Direct measurements of deep-brain and body-core temperature were performed on rats to determine the influence of cerebral blood flow (CBF) on brain temperature regulation under static and dynamic conditions. Static changes of CBF were achieved using different anesthetics (chloral hydrate, CH; alpha-chloralose, alphaCS; and isoflurane, IF) with alphaCS causing larger decreases in CBF than CH and IF; dynamic changes were achieved by inducing transient hypercapnia (5% CO(2) in 40% O(2) and 55% N(2)). Initial deep-brain/body-core temperature differentials were anesthetic-type dependent with the largest differential observed with rats under alphaCS anesthesia (ca. 2 degrees C). Hypercapnia induction raised rat brain temperature under all three anesthesia regimes, but by different anesthetic-dependent amounts correlated with the initial differentials--alphaCS anesthesia resulted in the largest brain temperature increase (0.32 +/- 0.08 degrees C), while CH and IF anesthesia lead to smaller increases (0.12 +/- 0.03 and 0.16 +/- 0.05 degrees C, respectively). The characteristic temperature transition time for the hypercapnia-induced temperature increase was 2-3 min under CH and IF anesthesia and approximately 4 min under alphaCS anesthesia. We conclude that both, the deep-brain/body-core temperature differential and the characteristic temperature transition time correlate with CBF: a lower CBF promotes higher deep-brain/body-core temperature differentials and, upon hypercapnia challenge, longer characteristic transition times to increased temperatures.

Cortical spreading depression (CSD), a transient neuronal and glial depolarization that propagates slowly across the cerebral cortex, is the putative electrophysiological event underlying migraine aura. It negatively impacts tissue injury during stroke, cerebral contusion and intracranial hemorrhage. Susceptibility to CSD has been assessed in several experimental animal models in vivo, such as after topical KCl application or cathodal stimulation. Various combinations of anesthetics and ambient conditions have been used by different laboratories making comparisons problematic and differences in data difficult to reconcile. We systematically studied CSD susceptibility comparing commonly used experimental anesthetics (isoflurane, alpha-chloralose, and urethane) with or without N(2)O or normobaric hyperoxia (100% O(2) inhalation). The frequency of evoked CSDs, and their propagation speed, duration, and amplitude were recorded during 2 h topical KCl (1 M) application. We found that N(2)O reduced CSD frequency when combined with isoflurane or urethane, but not alpha-chloralose; N(2)O also decreased CSD propagation speed and duration. Urethane anesthesia was associated with the highest CSD frequency that was comparable to pentobarbital. Inhalation of 100% O(2) did not alter CSD frequency, propagation speed or duration in combination with any of the anesthetics tested. Our data show anesthetic modulation of CSD susceptibility in an experimental model of human disease, underscoring the importance of proper study design for hypothesis testing as well as for comparing results between studies.

Understanding the link between the hemodynamic response and the underlying neuronal activity is important for interpreting functional magnetic resonance (fMRI) signals in human and animal studies. Simultaneous electrophysiological and functional imaging measurements provide a knowledge of information processing and communication in the brain with high spatial and temporal resolution. In this study, a range of neural and blood oxygenation level-dependent (BOLD) responses were elicited in the rat somatosensory cortex by changing the type of anesthesia (urethane or alpha-chloralose) and the electrical forepaw stimulus frequency (1–15 Hz). Duration of the stimulus was 30 s. Electrical local field potential and BOLD fMRI responses were recorded simultaneously. Under urethane anesthesia, integrated neural activity and BOLD responses increased with increasing stimulus frequency up to 11 Hz, after which both responses plateaued. In contrast, in alpha-chloralose-anesthetized rats both responses were measurable only at 1 and 3 Hz. Although neuronal and BOLD responses were nonlinear as a function of frequency over the 1 to 15 Hz stimulation range under both anesthetics, tight neural–hemodynamic coupling was observed independently of the anesthetic agent. Anesthetic agents influence neuronal activity in a different manner, but the relationship of neuronal activity and BOLD response remains the same.

Anticoagulant rodenticides are the largest group of poisons used to kill harmful rodents. Their fundamental mode of action consists in the inhibition of the vitamin K epoxide reductase, which causes blood-clotting alteration, ultimately leading to hemorrhagic events as the cause of death. In this study, we developed an UHPLC-MS-MS for the simultaneous determination of 10 anticoagulant hydroxycoumarine rodenticides, plus α-chloralose in human hair, with the scope of detecting potential trace of chronological poison exposure in clinical and forensic cases. The method was fully validated and applied to a case of intentional poisoning perpetrated by administration of difenacoum and α-chloralose to a 97-year-old woman, who was hospitalized because of severe symptoms, including drowsiness, convulsions, pallor and hematoma. Hair sample from the victim was segmentally analyzed. Difenacoum was detected in the proximal 3-cm hair segment at the concentration of 2.9 pg/mg. To our knowledge, this is the first study to report that exposure to difenacoum is detectable in real hair samples. The other target analyte found in the hair sample was α-chloralose, which was detected in the 0-3 cm segment at the concentration of 85 pg/mg. The two subsequent and consecutive segments (3-6 cm and 6-9 cm) showed only traces of difenacoum (below LOQ) and low but quantifiable concentrations of α-chloralose (29 and 6 pg/mg, respectively). Therefore, hair segmental analysis allowed us to conclude that the victim was repeatedly exposed to two poisons in the period corresponding to the first segment of hair.