Pentobarbital-anesthetized Animals Research Articles

Dietary Fish Oil Does Not Alter Glucose Tolerance in Conscious Rats

We examined the effect of dietary fish oil (MaxEPA) and sunflower seed oil on glucose tolerance in male Wistar rats. Semipurified diets containing 100 g oil/kg diet were administered for 30 d. The fish oil diet contained 26 g (n-3) fatty acids, 16 g eicosapentaenoic acid and 10.4 g docosahexaenoic acid/kg diet. Phospholipids from liver, pancreas, and pancreatic islets were enriched in eicosapentaenoic and docosahexaenoic acids by the fish oil diet. In unfed pentobarbital-anesthetized rats, both basal plasma insulin concentration and insulin responses to intravenous glucose were significantly lower for fish oil-fed rats although glucose responses were similar; however, incremental excursions in plasma insulin over the basal concentrations did not differ. Intravenous glucose tolerance was also examined in conscious unfed rats under minimal restraint. Responses of plasma glucose and insulin were similar for fish oil- and sunflower oil-fed groups. Furthermore, in another experiment, intravenous glucose tolerance tests were similar for conscious rats provided with either 100 g fish oil or corn oil/kg nonpurified diet. Thus, glucose-induced insulin secretion is lower in rats fed fish oil than in rats fed sunflower oil, when tests are conducted in pentobarbital-anesthetized animals but not when tests are performed in conscious rats; there was no effect on plasma glucose in either anesthetized or nonanesthetized rats. Therefore, substitution of (n-3) for (n-6) polyunsaturated fatty acids in tissue phospholipids does not alter plasma glucose or insulin in conscious male Wistar rats.

Pharmacological suppression of endogenous glucocorticoid synthesis attenuated blood pressure and heart rate response to acute restraint in Wistar rats.

Glucocorticoids (GCS) are known to modulate cardiovascular response during stress conditions. The present study was aimed to test the hypothesis that permissive and/or stimulating effect of GCs is essential for the maintenance of peripheral vascular resistance and for the adequate response of cardiovascular system to stressor exposure. The effects of acute pharmacological adrenalectomy (PhADX) on humoral and cardiovascular parameters were studied in adult Wistar rats under the basal conditions and during the acute restraint stress. Acute PhADX was performed by the administration of metyrapone and aminoglutethimide (100 mg/kg s.c. of each drug) resulting in a suppression of endogenous glucocorticoid synthesis. Blood pressure (BP), heart rate (HR) and core body temperature were measured using radiotelemetry. BP responses to administration of vasoactive agents were determined in pentobarbital-anesthetized animals. PhADX considerably attenuated stress-induced increase of BP, HR and core body temperature. PhADX did not abolish BP and HR lowering effects of ganglionic blocker pentolinium indicating preserved sympathetic function in PhADX rats. BP response to exogenous norepinephrine administration was attenuated in PhADX rats, suggesting reduced sensitivity of cardiovascular system. Suppression of corticosterone synthesis by PhADX increased basal plasma levels of ACTH, aldosterone and plasma renin activity in unstressed animals but there was no further increase of these hormones following stressor exposure. In conclusion, PhADX attenuated stress-induced rise of blood pressure, heart rate and core body temperature indicating an important permissive and/or stimulating role of glucocorticoids in the maintenance of the adequate response of cardiovascular system and thermoregulation to several stimuli including acute exposure to stressor.

The anti-inflammatory effects of methane*

Gastrointestinal methane generation has been demonstrated in various stress conditions, but it is not known whether nonasphyxiating amounts have any impact on the mammalian pathophysiology. We set out to characterize the effects of exogenous methane administration on the process of inflammatory events arising after reoxygenation in a large animal model of ischemia-reperfusion. A randomized, controlled in vivo animal study. A university research laboratory. Inbred beagle dogs (12.7 6 2 kg). Sodium pentobarbital-anesthetized animals were randomly assigned to sham-operated or ischemia-reperfusion groups, where superior mesenteric artery occlusion was maintained for 1 hr and the subsequent reperfusion was monitored for 3 hrs. For 5 mins before reperfusion, the animals were mechanically ventilated with normoxic artificial air with or without 2.5% methane. Biological responses to methane-oxygen respirations were defined in pilot rat studies and assay systems were used with xanthine oxidase and activated canine granulocytes to test the in vitro bioactivity potential of different gas concentrations. The macrohemodynamics and small intestinal pCO(2) gap changes were recorded and peripheral blood samples were taken for plasma nitrite/nitrate and myeloperoxidase analyses. Tissue superoxide and nitrotyrosine levels and myeloperoxidase activity changes were determined in intestinal biopsy samples; structural mucosal damage was measured by hematoxylin and eosin staining. Methane inhalation did not influence the macrohemodynamics but significantly reduced the magnitude of the tissue damage and the intestinal pCO(2) gap changes after reperfusion. Furthermore, the plasma and mucosal myeloperoxidase activity and the intestinal superoxide and nitrotyrosine levels were reduced, whereas the plasma nitrite/nitrate concentrations were increased. Additionally, methane effectively and specifically inhibited leukocyte activation in vitro. These data demonstrate the anti-inflammatory profile of methane. The study provides evidence that exogenous methane modulates leukocyte activation and affects key events of ischemia-reperfusion-induced oxidative and nitrosative stress and is therefore of potential therapeutic interest in inflammatory pathologies.

Severity of Locomotor and Cardiovascular Derangements after Experimental High-Thoracic Spinal Cord Injury Is Anesthesia Dependent in Rats

Anesthetics affect outcomes from central nervous system (CNS) injuries differently. This is the first study to show how two commonly used anesthetics affect continuously recorded hemodynamic parameters and locomotor recovery during a 2-week period after two levels of contusion spinal cord injury (SCI) in rats. We hypothesized that the level of cardiovascular depression and recovery of locomotor function would be dependent upon the anesthetic used during SCI. Thirty-two adult female rats were subjected to a sham, 25-mm or 50-mm SCI at T3-4 under pentobarbital or isoflurane anesthesia. Mean arterial pressure (MAP) and heart rate (HR) were telemetrically recorded before, during, and after SCI. Locomotor function recovered best in the 25-mm-injured isoflurane-anesthetized animals. There was no significant difference in locomotor recovery between the 25-mm-injured pentobarbital-anesthetized animals and the 50-mm-injured isoflurane-anesthetized animals. White matter sparing and extent of intermediolateral cell column loss appeared larger in animals anesthetized with pentobarbital, but this was not significant. There were no differential effects of anesthetics on HR and MAP before SCI, but recovery from anesthesia was significantly slower in pentobarbital-anesthetized animals. At the time of SCI, MAP was acutely elevated in the pentobarbital-anesthetized animals, whereas MAP decreased in the isoflurane-anesthetized animals. Hypotension occurred in the pentobarbital-anesthetized groups and in the 50-mm-injured isoflurane-anesthetized group. In pentobarbital-anesthetized animals, SCI resulted in acute elevation of HR, although HR remained low. Return of HR to baseline was much slower in the pentobarbital-anesthetized animals. Severe SCI at T3 produced significant chronic tachycardia that was injury severity dependent. Although some laboratories monitor blood pressure, HR, and other physiological variables during surgery for SCI, inherently few have monitored cardiovascular function during recovery. This study shows that anesthetics affect hemodynamic parameters differently, which in turn can affect functional outcome measures. This supports the need for a careful evaluation of cardiovascular and other physiological measures in experimental models of SCI. Choice of anesthetic should be an important consideration in experimental designs and data analyses.

Effects of colloid solutions on ischemia-reperfusion-induced periosteal microcirculatory and inflammatory reactions: Comparison of dextran, gelatin, and hydroxyethyl starch*

To compare the microhemodynamics and possible anti-inflammatory reactions of colloid resuscitation with 4% gelatin, 6% dextran, or 6% hydroxyethyl starch 130/0.4 solutions. A randomized control in vivo animal study in a university research laboratory. Adult male Wistar rats (280 +/- 20 g). Sodium pentobarbital-anesthetized animals were subjected to a 60-min complete hindlimb ischemia and a 180-min reperfusion. Volume resuscitation, either with a colloid (dextran, gelatin, or hydroxyethyl starch, 25 mL kg(-1) during 3 hr intravenously) or with lactated Ringer's solution, was initiated 10 min before reperfusion. Fluorescence intravital videomicroscopy was performed before ischemia and 30, 60, 120, and 180 min postresuscitation to quantify the tibial periosteal functional capillary density, the capillary red blood cell velocity changes, and leukocyte rolling and firm adherence in postcapillary venules. In a separate series blood samples were drawn to determine the release of soluble intercellular adhesion molecule-1 (enzyme-linked immunosorbent assay technique), and the surface expression of CD11b (flow cytometry) on peripheral blood granulocytes. Reperfusion significantly increased the leukocyte rolling and firm adhesion (by 2.6 and 7.1-fold, respectively), evoked marked decreases in periosteal functional capillary density and red blood cell velocity (56% and 39%, respectively), and increased the CD11b expression on the circulating leukocytes (by 85%). Hydroxyethyl starch, but not gelatin or dextran pretreatment, significantly inhibited the firm adherence of the leukocytes and reduced the elevated CD11b expression. Hydroxyethyl starch pretreatment also effectively attenuated the decreases in functional capillary density and red blood cell velocity, whereas gelatin and dextran did not improve the microhemodynamics. Finally, ischemia had no direct effect on the soluble intercellular adhesion molecule-1 levels, whereas gelatin treatment increased significantly this parameter. When compared with gelatin or dextran solutions, hydroxyethyl starch provided a therapeutic advantage in this setting by exerting an inhibitory effect on the ischemia-reperfusion-induced local and systemic leukocyte reactions and the postischemic periosteal microvascular dysfunction.

X-ray microanalysis of airway surface liquid in the mouse

The ionic composition of airway surface liquid (ASL) has been debated, and, in particular for the mouse, a wide range of values has been published. Two techniques were developed to measure the elemental composition of the ASL. X-ray microanalysis of ASL was carried out at low temperature on trachea removed from isoflurane-anesthetized animals and shock-frozen. In the second technique, dextran beads were placed on top of the epithelium of the trachea removed from pentobarbital-anesthetized animals, left to equilibrate with the ASL, dried, and subjected to X-ray microanalysis. Both techniques showed that mouse tracheal ASL has significantly lower concentrations of Na and Cl (approximately 60-80 mM) than serum. Differences between the two techniques were due to different sampling of mucus. CFTR(-/-) mice had significantly higher concentrations of Na and Cl in their ASL than age-matched controls. Pilocarpine or isoproterenol stimulation significantly reduced the ion concentrations in tracheal ASL. ASL was also collected with the dextran bead method from the nasal cavity in situ in pentobarbital-anesthetized animals. In control animals, the elemental composition of nasal fluid was similar to that of tracheal ASL. Pilocarpine stimulation caused a significant increase in Na, Cl, and K; stimulation with isoproterenol or phenylephrine caused a significant increase only in K. It is concluded that mouse ASL under unstimulated conditions is hypotonic, which may be related to the relative paucity of submucosal glands in the mouse trachea.

Propofol decreases diaphragmatic contractility in dogs.

Volatile anesthetics depress diaphragmatic muscle function; however, no data are available regarding the effect of propofol on diaphragmatic contractility. We therefore studied this effect in dogs. Pentobarbital-anesthetized animals were divided into three groups of 10 each. Group I received only maintenance fluid; Group II was infused with a subhypnotic dose of propofol (0.1-mg/kg initial dose plus 1.5-mg x kg(-1) x h(-1) maintenance dose); Group III was infused with an anesthetic dose of propofol (0.1-mg/kg initial dose plus 6.0-mg x kg(-1) x h(-1) maintenance dose). We assessed diaphragmatic contractility by transdiaphragmatic pressure (Pdi). With an infusion of propofol in Groups II and III, Pdi at low-frequency (20-Hz) stimulation decreased from the baseline values (P < 0.05), whereas Pdi at high-frequency (100-Hz) stimulation did not change. Compared with Group I, Pdi at 20-Hz stimulation decreased during propofol administration in Groups II and III (P < 0.05). The decrease in Pdi was more in Group III than in Group II (P < 0.05). We conclude that propofol is associated with a dose-related inhibitory effect on diaphragmatic contractility in dogs. Propofol is an effective IV anesthetic for the induction and maintenance of anesthesia. Subhypnotic and anesthetic doses of propofol decrease diaphragmatic contractility in dogs.

Pentobarbital-sensitive EDHF comediates ACh-induced arteriolar dilation in the hamster microcirculation.

It is unclear to what extent the endothelium-derived hyperpolarizing factor (EDHF) contributes to the control of microcirculatory blood flow in vivo. We analyzed, by intravital microscopy in hamster muscles, the potential role of EDHF along the vascular tree under stimulated (ACh) or basal conditions. Experiments were performed in conscious as well as anesthetized (pentobarbital, urethan) animals. Additionally, cellular effects of the potential EDHF were studied in isolated small arteries. In pentobarbital-anesthetized animals, treatment with Nomega-nitro-L-arginine (L-NNA; 30 micromol/l) and indomethacin (3 micromol/l) reduced the dilation in response to 10 micromol/l ACh from 60 +/- 6 to 20 +/- 4%. This nitric oxide/prostaglandin-independent dilation (NPID), which was of a similar magnitude in large and small arterioles, was abolished by potassium depolarization or charybdotoxin (ChTX, 1 micromol/l) but not by glibenclamide. In conscious animals, NPID amounted to 33 +/- 3%. The inhibitor of the P-450 monooxygenase 17-octadecynoic acid (ODYA) reduced NPID further to 9 +/- 4%. ChTX abolished the NPID and also reduced basal diameters (by -11 +/- 3%). The induction of anesthesia with pentobarbital reduced NPID (to 12 +/- 6%), whereas urethan anesthesia was without effect. Pentobarbital also reduced the ACh-induced hyperpolarization of vascular smooth muscle in isolated arteries, whereas ChTX abolished it. This study suggests that a considerable part of the ACh dilation in the microcirculation is mediated by EDHF, which also contributes to the control of basal tone in conscious animals. The direct inhibitory effect of pentobarbital and ODYA supports the idea that "microcirculatory" EDHF is a product of the cytochrome P-450 pathway. The role of EDHF might be underestimated in pentobarbital-anesthetized animals.

The Correlation of the Weakening Effect on Gastric Mucosal Integrity by 5-HT With Neutrophil Activation

The effects of 5-hydroxytryptamine (5-HT) on ethanol-induced gastric mucosal damage and on epithelial and vascular integrity were investigated. Male Sprague–Dawley rats were administered with 5-HT (5 or 10 mg/kg, IP) 30 min prior to the challenge with ethanol (40% v/v, 10 ml/kg, PO). 5-HT dose dependently aggravated ethanol-induced injury in the gastric mucosa. Both xanthine oxidase (XO) and myeloperoxidase (MPO) activities in the mucosa were significantly increased with the high dose of 5-HT, which also potentiated the elevation of these enzyme activities by ethanol. However, the mucosal superoxide dismutase activity was left unaltered. In neutropenic (antineutrophil serum-treated) animals, the ethanol-induced gastric mucosal injury was significantly ameliorated, with or without the pretreatment of 5-HT (10 mg/kg). In addition, the effect of 5-HT on the activity of MPO, but not of XO, was also attenuated in these animals. In the ex vivo gastric chamber study on pentobarbital-anesthetized animals, volume of gastric secretion was significantly decreased in the 5-HT–treated groups, with further reduction after ethanol incubation. Transmucosal potential difference (PD) was significantly reduced in 5-HT–treated rats, which also potentiated the ethanol-induced drop in PD. Nevertheless, 5-HT dose dependently increased mucosal vascular permeability and further enhanced during ethanol incubation. These findings suggest that 5-HT adversely affects the defense mechanisms of the gastric mucosa by reducing the secretory function of the mucosal cells and to weaken the epithelial and vascular integrity. Neutrophil activation appears to be responsible for the detrimental effects of 5-HT partly through the elevation in MPO activity. The increase in mucosal XO activity by 5-HT may induce free radical production and possibly modulate the ulcerogenic processes.

Preferential reduction in vascular responses to endothelin-1 in rats with streptozotocin-induced diabetes.

The effects of vasodepressor (acetylcholine and bradykinin) and pressor [electrical stimulation of the spinal sympathetic outflow, norepinephrine and endothelin-1 (ET-1)] stimuli were determined in rats with 2- and 5-week untreated streptozotocin-induced diabetes (blood glucose 400 and >500 mg/dl, respectively). In pentobarbital-anesthetized animals, the hypotensive response to an intravenous dose of acetylcholine or bradykinin was unaffected in animals treated for 2 weeks with streptozotocin but was significantly reduced (22% and 48%, respectively) after 5 weeks. However, the pressor responses to ET-1 were significantly decreased in animals that had been given streptozotocin 2 (38%) and 5 (45%) weeks previously. In contrast, the vasoconstrictor effects of electrical stimulation of the spinal cord outflow and norepinephrine were significantly inhibited (47% and 30%, respectively) at 5 weeks, but not at 2 weeks, after streptozotocin administration. These results indicate that, in untreated streptozotocin diabetes, a substantial impairment of vascular reactivity to ET-1 appears more rapidly than inhibition of the pressor responses to endogenous and exogenous norepinephrine or to vasodilator substances that require integrity of vascular endothelial cell function to produce their normal effects.

The influence of angiotensin II-induced increase in aortic wall mass on compliance in rats in vivo.

This study was undertaken to examine the effects of angiotensin II-induced structural changes in the aortic wall on the dynamic mechanical properties of the vessel in the rat. Wistar rats were infused s.c. with 250 ng/kg/min angiotensin II (Ang II) for 14 days (ANG). Both ANG and control rats (CON) were equipped with an arterial catheter for measurement of arterial blood pressure. Thoracic aorta diameter, compliance coefficient (CC), and distensibility coefficient (DC) were determined non-invasively in pentobarbital-anesthetized animals using a B-mode imager attached to a vessel wall tracking system. After sacrifice, medial cross-sectional area (CSA), and elastin and collagen densities were determined by morphometry on cross-sections. Media thickness (Mt) and wall-to-lumen ratio (W/L) were subsequently calculated. Ang II infusion significantly increased mean arterial blood pressure in conscious rats (122 +/- 3 mmHg CON vs. 157 +/- 4 mmHg ANG). This was normalized when the rats were anesthetized, thus making it possible to determine CC and DC under isobaric conditions where the diastolic diameters were also similar. Two-week infusion of Ang II induced a significant increase in CSA from 0.48 +/- 0.02 mm2 in CON to 0.61 +/- 0.03 mm2. Mt and W/L were likewise increased, but collagen and elastin densities remained unchanged. CC and DC were not effected by this increase in aortic wall mass (CC: 0.143 +/- 0.009 CON, 0.147 +/- 0.014 mm2/kPa ANG; DC: 0.052 +/- 0.005 CON, 0.051 +/- 0.004 kPa-1 ANG). An increase in aortic wall mass resulting from chronic infusion of angiotensin II does not alter the dynamic compliance of the vessel under isobaric conditions.

Effect of nitric oxide synthase inhibition on myocardial contractility in anesthetized normal and endotoxemic dogs.

Nitric oxide (NO) produced by the induced NO synthase (NOS) enzyme has been implicated in the mechanisms of the circulatory changes that occur in the later stages of sepsis. As NO produced by the constitutive form of the enzyme is known to play a role in the regulation of normal circulation, we have performed a series of experiments to study the early circulatory effects of inhibition of NOS in a hyperdynamic endotoxemic dog model. Pentobarbital-anesthetized animals were used. Cardiac output (CO) was measured by thermodilution. Myocardial contractility (MC) was estimated from the slope of the left ventricular end-systolic pressure-diameter relationship obtained from sonomicrometer- and catheter-tip manometer signals in closed chest animals. All animals received a 15 mL/kg/h infusion of Ringer's lactate. A hyperdynamic response was elicited by a 2 h infusion of a total dose of 5.3 micrograms/kg Escherichia coli O55:B5 endotoxin (ETX). CO increased initially by about 25%, and total peripheral resistance decreased by 35%. These changes subsided in 60-90 min, after which a sustained decrease in CO occurred. MC elevated transiently by 25% after the first 30 min of ETX infusion, then decreased gradually below the control level. Administration of 2 mg/kg of the NOS inhibitor N-nitro-L-arginine (NNA) between the 45th and 55th min of the ETX infusion increased MC to the level in the control group, but accelerated the decline of the initially increased CO and caused a sustained increase in total peripheral resistance to about 50% above the control level. In normal (nonendotoxin treated) dogs, NNA also caused a similar increase in MC which, however, lasted at least 3 h. Left ventricular diameter increased in the NNA-treated groups. This increase also occurred in the endotoxin-only group but with a delay of about 2.5 h. Our results demonstrate the participation of constitutive NOS-produced NO in the early hyperdynamic response of endotoxemia. Suppression of NO is associated with increased myocardial contractility. NNA treatment may be favorable for the restoration of depressed cardiac contractility during endotoxemia, but this treatment is probably detrimental for the compensatory systemic flow (CO) increase.

Cardiovascular effects of (-)-11-OH-delta 8-tetrahydrocannabinol-dimethylheptyl in rats.

The effects of the stereochemically pure psychoactive cannabinoid (-)-11-OH-delta 8-tetrahydrocannabinol-dimethylheptyl (HU-210) on blood pressure (BP) and heart rate (HR) were determined in rats. In pentobarbital-anesthetized animals, the compound produced dose-related, long-lasting hypotension and bradycardia at doses between 10 and 1,000 micrograms/kg. BP began to decrease immediately after drug administration, and in no case was an initial pressor response observed. Previous vagotomy (VX) or pretreatment with 6-hydroxydopamine (6-OHDA) did not affect hypotension. Bradycardia was inhibited by VX, but only 60 min after administration of HU-210; it was enhanced by 6-OHDA. The cannabinoid blocked reflex bradycardia induced by phenylephrine (PE). HU-210 also decreased BP and HR in conscious rats. Hypotension lasted 2 h, whereas bradycardia was still present 8 h after drug administration. HU-210 thus shares with delta 9-tetrahydrocannabinol (THC) the ability to decrease BP and HR, but is 5-10 times more potent than the natural compound. Its lack of an initial pressor effect, such as that described for THC, could be related to its specificity for the type-1 cannabinoid (CB1) receptor. Hypotension and bradycardia after HU-210 administration are not due to sympathetic withdrawal. Enhanced parasympathetic tone is involved in bradycardia only at a late stage of the response.

Comparison of response properties of neurons in the inferior colliculus of guinea pigs under different anesthetics.

Responses of the inferior colliculus (IC) neurons to acoustical stimulation were recorded in anesthetized guinea pigs. Three kinds of anesthesia were used: (1) urethane (8 ml/kg b.w. of 20 per cent solution i.p.); (2) ketamine-xylazine combination (1 ml/kg b.w. of mixture 2:1); and (3) pentobarbital (25 mg/kg i.p.) combined with intramuscular injection of fentanyl (0.5 ml) and droperidol (1 ml). The frequency tuning of neurons evaluated on the basis of Q10 values and the composite neural audiogram represented by points of lowest thresholds of individual IC neurons were similar for guinea pigs treated with any of the anesthetics. The number of spontaneously active IC neurons was significantly larger with ketamine than with urethane or pentobarbital. The response latencies to tone bursts at characteristic frequency (CF) were shortest in animals anesthetized with pentobarbital. Whereas with ketamine and urethane many neurons were recorded in which response latencies were longer than 40 ms, in pentobarbital-anesthetized animals the latencies usually did not exceed 25 ms. The occurrence of neurons with an onset type of response was significantly larger with pentobarbital than with the other two anesthetics. In ketamine and urethane anesthesia, thresholds of units with sustained response were significantly lower than thresholds of units with onset response. Our results suggest that in experiments where the level of spontaneous activity, latency and type of responses were important parameters, the kind of anesthesia should be taken into account.

Induction of heterosynaptic and homosynaptic LTD in hippocampal sub-regions in vivo

Establishing the conditions under which long-term depression (LTD) can be induced in vitro remains an important empirical issue. While heterosynaptic LTD is readily elicited in the hippocampus in vivo, the induction of homosynaptic LTD has been more problematic, both in CA1 and in the dentate gyrus. Our work has demonstrated, however, that 900 pulses at 1 Hz will reliably induce LTD in both area CA1 and the dentate gyrus of pentobarbital-anesthetized animals, when the stimulating and recording electrodes are in close proximity to each other.

Effect of acetylcholine on corticotropin-releasing factor gene expression in the hypothalamic paraventricular nucleus of conscious rats.

To examine the physiological role of cholinergic input in the regulation of CRF neurons in the paraventricular nucleus (PVN) of the hypothalamus, acetylcholine (ACh) was microinjected bilaterally into the dorsolateral border of the PVN of conscious rats. Changes in the levels of POMC messenger RNA (mRNA) in the anterior pituitary, CRF mRNA in hypothalamic tissue containing the PVN, and plasma ACTH were assessed. Plasma ACTH concentrations increased in a dose-dependent manner after ACh injection (1-100 pmol/side), reaching a peak 30 min after ACh injection and returning to baseline within 120 min. The POMC mRNA level in the anterior pituitary and the hypothalamic CRF mRNA level increased in a dose-dependent manner 120 min after ACh (0.1-10 pmol/side) injection. Intracerebroventricular pretreatment with atropine completely abolished the ACh-induced increase in plasma ACTH concentrations, whereas pretreatment with hexamethonium was without significant effect. The intracerebroventricular injection of ACh also increased plasma ACTH concentrations in a dose-dependent manner in conscious rats, but not in pentobarbital-anesthetized animals. Thus, cholinergic hypothalamic input stimulates CRF gene expression in the PVN and CRF secretion into the portal circulation under physiological conditions. The use of conscious animals is essential in elucidating the physiological roles of neurotransmitters and other modulators regulating CRF neurons.

Effect of acetylcholine on corticotropin-releasing factor gene expression in the hypothalamic paraventricular nucleus of conscious rats

To examine the physiological role of cholinergic input in the regulation of CRF neurons in the paraventricular nucleus (PVN) of the hypothalamus, acetylcholine (ACh) was microinjected bilaterally into the dorsolateral border of the PVN of conscious rats. Changes in the levels of POMC messenger RNA (mRNA) in the anterior pituitary, CRF mRNA in hypothalamic tissue containing the PVN, and plasma ACTH were assessed. Plasma ACTH concentrations increased in a dose-dependent manner after ACh injection (1-100 pmol/side), reaching a peak 30 min after ACh injection and returning to baseline within 120 min. The POMC mRNA level in the anterior pituitary and the hypothalamic CRF mRNA level increased in a dose-dependent manner 120 min after ACh (0.1-10 pmol/side) injection. Intracerebroventricular pretreatment with atropine completely abolished the ACh-induced increase in plasma ACTH concentrations, whereas pretreatment with hexamethonium was without significant effect. The intracerebroventricular injection of ACh also increased plasma ACTH concentrations in a dose-dependent manner in conscious rats, but not in pentobarbital-anesthetized animals. Thus, cholinergic hypothalamic input stimulates CRF gene expression in the PVN and CRF secretion into the portal circulation under physiological conditions. The use of conscious animals is essential in elucidating the physiological roles of neurotransmitters and other modulators regulating CRF neurons.

Antifibrillary Action of Class I-IV Antiarrhythmic Agents in the Model of Ventricular Fibrillation Threshold of Anesthetized Guinea Pigs

We compared the effects of class I-IV antiarrhythmic agents on the ventricular fibrillation threshold (VFT) induced by electrical stimulation directly on the myocardium in anesthetized, open-chest guinea pigs. VFT was assessed by determining the intensity (mA) of electrical current required to induce ventricular fibrillation (VF) and is expressed as a percentage change of the baseline premedication value. The following antiarrhythmic agents or their solvent were administered intravenously (i.v.) to pentobarbital-anesthetized animals (n = 6-12 per group): class I antiarrhythmic agent encainide (1.5 mg/kg); class II antiarrhythmic agents atenolol (2.5 mg/kg), metoprolol (2.5 mg/kg), and nebivolol (2.5 mg/kg); class III antiarrhythmic agents dofetilide (0.08 mg/kg), terikalant (0.04 mg/kg), and DL-sotalolol (10 mg/kg); and class IV antiarrhythmic agent verapamil (0.16 mg/kg). The antiarrhythmic compounds or their solvents resulted in the following changes in the VFT at 15 min after treatment: saline control, 1 +/- 14% (mean +/- SEM) from its baseline value; 10% hydroxypropyl-beta-cyclodextrine (CD), 4 +/- 13%; encainide, 183 +/- 46% (p < 0.05 vs. saline); atenolol, 66 +/- 23% (p > 0.05 vs. saline); metoprolol, 89 +/- 25% (p > 0.05 vs. saline); nebivolol, 224 +/- 58% (p < 0.05 vs. 10% CD); DL-sotalol, 485 +/- 119% (p < 0.05 vs. saline); dofetilide, 357 +/- 69% (p < 0.05 vs. saline); terikalant, 487 +/- 183% (p < 0.05 vs. saline), and verapamil, -17 +/- 21% (p > 0.05 vs. saline). At the doses used, all compounds significantly reduced heart rate (HR).(ABSTRACT TRUNCATED AT 250 WORDS)

Basal and reactive plasma catecholamine levels under stress and anesthesia in rabbits

The present study was undertaken to determine if electrical stimulation of the hypothalamic defense area (HDA) and baroreflex activation elicited by head up body tilt produced changes in plasma catecholamine (CA) levels in anesthetized rabbits. We also compared the effects of two anesthetics, isoflurane and sodium pentobarbital, upon basal and reactive CA levels, and upon autonomic reactivity. HDA stimulation was found to produce significant increases in plasma norepinephrine (NE) levels but not epinephrine (E) levels. Passive tilt was found to produce statistically significant increases in NE levels for both anesthetics used and a significant increase in E levels for animals anesthetized with isoflurane. Basal and reactive measurements provided evidence that pentobarbital has a more suppressive effect upon the autonomic nervous system than isoflurane: (a) Basal NE levels were significantly lower in pentobarbital-anesthetized animals than in isoflurane-anesthetized animals; and (b) Baroreceptor sensitivity to a passive tilt stressor was significantly higher for animals anesthetized with isoflurane than for animals anesthetized with pentobarbital.

Interaction of Anesthetics and Catecholamines on Conduction in the Canine His--Purkinje System

The findings in papillary muscles that epinephrine facilitates conduction at Purkinje fiber-muscle junctions and in the endocardium are consistent with older observations that activation of myocardial beta-adrenergic receptors speeds conduction and activation in the heart and thereby increases the synergy of contraction (46,47). The cellular mechanism underlying this action is probably increased cell-to-cell coupling between muscle fibers secondary to elevation of cyclic AMP (19,48). However, the findings that epinephrine alone or with halothane transiently slows conduction in the Purkinje layer while simultaneously improving conduction across Purkinje-muscle junctions and in the endocardium may represent proarrhythmic actions. These actions could facilitate arrhythmogenesis by transiently increasing regional differences of activation and repolarization times in the conduction system and myocardium and thereby increasing vulnerability to induction of reentry by premature impulses. Such a proarrhythmic effect could explain an older observation that low-dose norepinephrine infusions decrease the threshold for induction of fibrillation by two premature beats in pentobarbital-anesthetized animals (49). The cellular basis underlying the different responses of Purkinje fibers and the endocardial muscle layer to catecholamines, in which velocity decreased and increased, respectively, is not known. Our working hypothesis to explain this action in canine Purkinje fibers is a mechanism involving activation of WB4101-sensitive alpha 1-adrenoceptor, G-protein coupling to phospholipase C and the generation of DAG and IP3 leading to modulation of cell-to-cell coupling, which is potentiated in the presence of partial uncoupling by halothane. The different responses of Purkinje and myocardial fibers are speculated to result from differences in the relative density of this subtype of alpha 1-adrenoceptor, differences in the subcellular effector coupling mechanisms, or differences in the specific connexin proteins forming gap junctions between Purkinje and myocardial fibers (50).