Muscarinic Blocker Research Articles

Abstract 66: Early-Life Sodium Restriction Programs Salt-Sensitivity of Autonomics and Blood Pressure in Adult C57BL/6J Mice

Extremely premature birth is a risk factor for sodium (Na) depletion due to renal immaturity and urine Na losses. We have proposed that Na depletion in preterm infants mechanistically contributes to their increased risk for cardiovascular and metabolic disease observed in adulthood, potentially due to changes during hypothalamic development. In humans, the hypothalamus matures between 20-25 weeks of gestation; in mice, this developmental window occurs between postnatal day (PD) 21 and PD42. Previously, we demonstrated in mice that dietary Na restriction from PD21 to PD42 induced long-lasting increases in energy expenditure via exaggerated autonomic activity. Therefore, we hypothesized that restricting dietary Na from PD21 to PD42 would similarly induce programmed changes in cardiovascular control via changes in autonomic and neurohypophyseal functions. Male C57BL/6J mice were provided a customized low (0.04%, n=7) or supplemented (0.30%, n=8) Na version of the soy protein-free Teklad 2920x diet from PD21 to PD42, after which they were returned to the standard (0.15% Na) 2920x diet. Radiotelemetric blood pressure (BP) transducers were implanted at 7-8 weeks of age. At 10 weeks of age, no differences in BP or heart rate (HR) were noted between groups. All animals were switched to a high (1%) Na diet at 12 weeks of age. At 16 weeks of age, systolic BP was not altered in the 0.30% Na group but significantly increased in the 0.04% Na group compared to the week 10 baseline (24h avg: +6±2 mmHg, p<0.05). Ganglionic block with chlorisondamine or β-adrenergic block with propranolol reduced heart rate (HR) to a greater extent (each p<0.05) in the 0.04% group. Conversely, muscarinic block with atropine resulted in a smaller increase in HR in this group (p<0.05). Urine elimination rates of aldosterone (0.30%: 1.96±0.44 vs 0.04%: 2.92±0.064 ng/d) and copeptin (0.30%: 0.16±0.03 vs. 0.04%: 0.14±0.02 ng/d) were similar between groups. Finally, at 18 weeks of age, all mice received the angiotensin AT 1 R antagonist losartan in drinking water. This caused a greater BP reduction in the 0.04% group at 20 weeks (24h avg: -12±2 vs -19±2 mmHg, p<0.05). Together, these data indicate that dietary Na restriction during hypothalamic development programs increased salt sensitivity of BP via a mechanism involving AT 1 R and characterized by increased sympathetic yet decreased parasympathetic activity.

NEUROBEHAVIOURAL AND ANALGESIC EFFECT OF Ocimum gratissimum LINN. LEAVES ESSENTIAL OIL IN Wistar albino MICE

Studies have shown that pain relieving medications may be neuroprotective. Ocimum gratissimum Linn. that is widely used in traditional medicine for debility and many other illnesses neuropharmacologically related has not been fully explored. Aim. This study was designed to investigate the safety of intake, neurobehavioral and analgesic effects of the Essential Oil of Ocimum gratissimum Linn leaves (EOOG) in mice. Methods. Acute toxicity of EOOG was determined following standard method while the neurobehavioural properties were assessed using the open field for Novelty-Induced Rearing (NIR), Novelty-Induced Grooming (NIG) and locomotor activity in mice. The hole board apparatus was used for the frequency of head dips. The Y-maze was used for short- working memory. Mechanistic studies were conducted with Atropine (muscarinic blocker, 0.5 mg/kg), Propanolol (non-selective ß-adrenoceptor blocker, 0.2 mg/kg), Haloperidol (dopamine receptor blocker, 0.2 mg/kg), Cyproheptadine (Serotoninergic antagonist, 0.5 mg/kg) and Yohimbine (ά-2 adrenergic blocker, 1 mg/kg). The analgesic activity of Ocimum gratissimum was investigated using acetic acid writhing test and thermally-induced pain. Results. The median lethal dose (LD50) of Ocimum gratissimum was 2449 mg/kg. The EOOG significantly reduced novelty-induced behaviour in a dose-dependent manner. The exploratory activity of animals treated with the EOOG was observed to decrease non-dose dependently with the highest dose (40 mg/kg) showing no activity on the hole board apparatus. The EOOG produced a significant reduction in locomotor activity in all the doses in a non-dose dependent manner but at the lowest dose. In the Y-maze, EOOG did not produce any significant effect on working memory as the percentage alternation produced was not significantly different from the control. The EOOG in hot plate analgesic assay showed increased reaction time suggesting central nervous system analgesic property. Conclusions. The results of the investigation showed that EOOG might possess sedative properties due to its ability to inhibit NIR and NIG, head dips, and locomotor activity. Furthermore, the inhibition of nociception marked in this research advocates antinociceptive activity which might be through the peripheral or central opioid receptor..

The cAMP-phosphodiesterase 4 (PDE4) controls β-adrenoceptor- and CFTR-dependent saliva secretion in mice

Saliva, while often taken for granted, is indispensable for oral health and overall well-being, as inferred from the significant impairments suffered by patients with salivary gland dysfunction. Here, we show that treatment with several structurally distinct PAN-PDE4 inhibitors, but not a PDE3 inhibitor, induces saliva secretion in mice, indicating it is a class-effect of PDE4 inhibitors. In anesthetized mice, while neuronal regulations are suppressed, PDE4 inhibition potentiates a β-adrenoceptor-induced salivation, that is ablated by the β-blocker Propranolol and is absent from homozygous ΔF508-CFTR mice lacking functional CFTR. These data suggest that PDE4 acts within salivary glands to gate saliva secretion that is contingent upon the cAMP/PKA-dependent activation of CFTR. Indeed, PDE4 contributes the majority of total cAMP-hydrolytic capacity in submandibular-, sublingual-, and parotid glands, the three major salivary glands of the mouse. In awake mice, PDE4 inhibitor-induced salivation is reduced by CFTR deficiency or β-blockers, but also by the muscarinic blocker Atropine, suggesting an additional, central/neuronal mechanism of PDE4 inhibitor action. The PDE4 family comprises four subtypes, PDE4A-D. Ablation of PDE4D, but not PDE4A-C, produced a minor effect on saliva secretion, implying that while PDE4D may play a predominant role, PDE4 inhibitor-induced salivation results from the concurrent inactivation of multiple (at least two) PDE4 subtypes. Taken together, our data reveal a critical role for PDE4/PDE4D in controlling CFTR function in an in vivo model and in inducing salivation, hinting at a therapeutic potential of PDE4 inhibition for cystic fibrosis and conditions associated with xerostomia.

Parasympathetic brainstem origin sites of triggered external and internal carotid vasodilation are not distributed topographically in the rat

Cranial parasympathetic activation produces vasodilation in the head and neck region, but little is known about its central and peripheral mechanisms. This study was conducted to examine whether external and internal carotid-vasodilation origin sites triggered by chemical stimulation are distributed topographically in the parasympathetic brainstems of anesthetized rats, and to examine the effects of peripheral receptors on vasodilation. Microinjection of the neuromodulator candidate l-cysteine revealed that external and internal carotid vasodilation–triggering sites were distributed non-topographically along the full extent of the parasympathetic parvocellular reticular formation (PcRt). Intravenous injection of a muscarinic blocker and a nitric oxide synthase inhibitor abolished external carotid vasodilation, suggesting the peripheral involvement of muscarinic and nitric oxide receptors. Further work is needed to fully understand the PcRt mechanisms underlying timely and appropriate vasodilation to support various cranial functions.

Biphasic changes in spontaneous cardiovagal baroreflex sensitivity during passive hyperthermia

Successful adaptation to passive hyperthermia requires continual adjustment of circulation, which is mediated mainly by the autonomic nervous system. The goal of this study was to explore the alterations in spontaneous cardiovagal baroreflex sensitivity (BRS) during exposure to a hot environment. To continuously follow changes in core body temperature (Tc), haemodynamics, and BRS, male Wistar-Kyoto rats were implanted with telemetric transmitters. BRS at an ambient temperature of 23 °C was not steady but oscillated with a maximum power in the range of 0.02–0.2 Hz. Exposure to hot air immediately shifted the distribution of BRS to higher values, although Tc remained unchanged (37.2 (0.3) °C), and the average BRS changed from 1.3 (0.3) to 3 (1.4) ms.mmHg−1, p < 0.0001. The degree of initial cardiovagal baroreflex sensitization explained 57% of the variability in the time to the onset of arterial pressure decline (p = 0.0114). With an increasing Tc (>38.8 (0.6) °C), BRS non-linearly declined, but haemodynamic parameters remained stable even above a Tc of 42 °C when the cardiovagal baroreflex was virtually non-operative. Abrupt full desensitization of the cardiovagal baroreflex with a muscarinic blocker did not induce arterial pressure decline. Our data indicate that a progressive decrease in BRS during passive hyperthermia does not induce haemodynamic instability. The positive association between initial cardiovagal baroreflex sensitization and the time to the onset of arterial pressure decline may reflect the potential protective role of parasympathetic activation during exposure to a hot environment.

Effect of chronic exercise on myocardial electrophysiological heterogeneity and stability. Role of intrinsic cholinergic neurons: A study in the isolated rabbit heart.

A study has been made of the effect of chronic exercise on myocardial electrophysiological heterogeneity and stability, as well as of the role of cholinergic neurons in these changes. Determinations in hearts from untrained and trained rabbits on a treadmill were performed. The hearts were isolated and perfused. A pacing electrode and a recording multielectrode were located in the left ventricle. The parameters determined during induced VF, before and after atropine (1μM), were: fibrillatory cycle length (VV), ventricular functional refractory period (FRPVF), normalized energy (NE) of the fibrillatory signal and its coefficient of variation (CV), and electrical ventricular activation complexity, as an approach to myocardial heterogeneity and stability. The VV interval was longer in the trained group than in the control group both prior to atropine (78±10 vs. 68±10 ms) and after atropine (76±8 vs. 67±10 ms). Likewise, FRPVF was longer in the trained group than in the control group both prior to and after atropine (53±8 vs. 42±7 ms and 50±6 vs. 40±6 ms, respectively), and atropine did not modify FRPVF. The CV of FRPVF was lower in the trained group than in the control group prior to atropine (12.5±1.5% vs. 15.1±3.8%) and, decreased after atropine (15.1±3.8% vs. 12.2±2.4%) in the control group. The trained group showed higher NE values before (0.40±0.04 vs. 0.36±0.05) and after atropine (0.37±0.04 vs. 0.34±0.06; p = 0.08). Training decreased the CV of NE both before (23.3±2% vs. 25.2±4%; p = 0.08) and after parasympathetic blockade (22.6±1% vs. 26.1±5%). Cholinergic blockade did not modify these parameters within the control and trained groups. Activation complexity was lower in the trained than in the control animals before atropine (34±8 vs. 41±5), and increased after atropine in the control group (41±5 vs. 48±9, respectively). Thus, training decreases the intrinsic heterogeneity of the myocardium, increases electrophysiological stability, and prevents some modifications due to muscarinic block.

Tone identification behavior in Rattus norvegicus: muscarinic receptor blockage lowers responsiveness in nontarget selective neurons, while nicotinic receptor blockage selectively lowers target responses.

Learning to associate a stimulus with reinforcement causes plasticity in primary sensory cortex. Neural activity caused by the associated stimulus is paired with reinforcement, but population analyses have not found a selective increase in response to that stimulus. Responses to other stimuli increase as much as, or more than, responses to the associated stimulus. Here, we applied population analysis at a new time point and additionally evaluated whether cholinergic receptor blockers interacted with the plastic changes in cortex. Three days of tone identification behavior caused responsiveness to increase broadly across primary auditory cortex, and target responses strengthened less than overall responsiveness. In pharmacology studies, behaviorally impairing doses of selective acetylcholine receptor blockers were administered during behavior. Neural responses were evaluated on the following day, while the blockers were absent. The muscarinic group, blocked by scopolamine, showed lower responsiveness and an increased response to the tone identification target that exceeded both the 3-day control group and task-naïve controls. Also, a selective increase in the late phase of the response to the tone identification stimulus emerged. Nicotinic receptor antagonism, with mecamylamine, more modestly lowered responses the following day and lowered target responses more than overall responses. Control acute studies demonstrated the muscarinic block did not acutely alter response rates, but the nicotinic block did. These results lead to the hypothesis that the decrease in the proportion of the population spiking response that is selective for the target may be explained by the balance between effects modulated by muscarinic and nicotinic receptors.

The Organophosphate Paraoxon and Its Antidote Obidoxime Inhibit Thrombin Activity and Affect Coagulation In Vitro.

Organophosphates (OPs) are potentially able to affect serine proteases by reacting with their active site. The potential effects of OPs on coagulation factors such as thrombin and on coagulation tests have been only partially characterized and potential interactions with OPs antidotes such as oximes and muscarinic blockers have not been addressed. In the current study, we investigated the in vitro interactions between coagulation, thrombin, the OP paraoxon, and its antidotes obidoxime and atropine. The effects of these substances on thrombin activity were measured in a fluorescent substrate and on coagulation by standard tests. Both paraoxon and obidoxime but not atropine significantly inhibited thrombin activity, and prolonged prothrombin time, thrombin time, and partial thromboplastin time. When paraoxon and obidoxime were combined, a significant synergistic effect was found on both thrombin activity and coagulation tests. In conclusion, paraoxon and obidoxime affect thrombin activity and consequently alter the function of the coagulation system. Similar interactions may be clinically relevant for coagulation pathways in the blood and possibly in the brain.

A functional study on small intestinal smooth muscles in jejunal atresia.

Aim:The present study was aimed to assess the contractile status of neonatal small intestinal smooth muscle of dilated pre-atretic part of intestinal atresia to resolve debatable issues related to mechanisms of persistent dysmotility after surgical repair.Materials and Methods:A total of 34 longitudinally sectioned strips were prepared from pre-atretic dilated part of freshly excised 8 jejunal atresia type III a cases. Spontaneous as well as acetylcholine- and histamine-induced contractions were recorded in vitro by using organ bath preparations. Chemically evoked contractions were further evaluated after application of atropine (muscarinic blocker), pheniramine (H1 blocker), and lignocaine (neuronal blocker) to ascertain receptors and neuronal involvement. Histological examinations of strips were made by using Masson trichrome stain to assess the fibrotic changes.Results:All 34 strips, except four showed spontaneous contractions with mean frequency and amplitude of 5.49 ± 0.26/min and 24.41 ± 5.26 g/g wet tissue respectively. The response to ACh was nearly twice as compared to histamine for equimolar concentrations (100 μM). ACh (100 μM) induced contractions were attenuated (by 60%) by atropine. Histamine (100 μM)-induced contractions was blocked by pheniramine (0.32 μM) and lignocaine (4 μM) by 74% and 78%, respectively. Histopathological examination showed varying degree of fibrotic changes in muscle layers.Conclusions:Pre-atretic dilated part of jejunal atresia retains functional activity but with definitive histopathologic abnormalities. It is suggested that excision of a length of pre-atretic part and early stimulation of peristalsis by locally acting cholinomimetic or H1 agonist may help in reducing postoperative motility problems in atresia patients.

A new potassium ion current induced by stimulation of M2-cholinoreceptors in fish atrial myocytes

A novel potassium ion current induced by muscarinic stimulation (IKACh2) is characterized in atrial cardiomyocytes of teleost fishes (crucian carp, Carassius carassius; rainbow trout, Oncorhynchus mykiss) by means of the whole-cell patch-clamp technique. The current is elicited in atrial, but not ventricular, cells by application of carbamylcholine (CCh) in moderate to high concentrations (10(-7)-10(-4) mol l(-1)). It can be distinguished from the classic IKACh, activated by the βγ-subunit of the Gi-protein, because of its low sensitivity to Ba(2+) ions and distinct current-voltage relationship with a very small inward current component. Ni(2+) ions (5 mmol l(-1)) and KB-R7943 (10(-5) mol l(-1)), non-selective blockers of the sodium-calcium exchange current (INCX), strongly reduced and completely abolished, respectively, the IKACh2. Therefore, IKACh2 was initially regarded as a CCh-induced outward component of the INCX. However, the current is not affected by either exclusion of intracellular Na(+) or extracellular Ca(2+), but is completely abolished by intracellular perfusion with K(+)-free solution. Atropine (10(-6) mol l(-1)), a non-selective muscarinic blocker, completely eliminated the IKACh2. A selective antagonist of M2 cholinoreceptors, AF-DX 116 (2×10(-7) mol l(-1)) and an M3 antagonist, 4-DAMP (10(-9) mol l(-1)), decreased IKACh2 by 84.4% and 16.6%, respectively. Pertussis toxin, which irreversibly inhibits Gi-protein coupled to M2 receptors, reduced the current by 95%, when applied into the pipette solution. It is concluded that IKACh2, induced by stimulation of M2 cholinoceptors and subsequent Gi-protein activation, represents a new molecular target for the cardiac parasympathetic innervation.

Vagal modulation of cardiac ventricular arrhythmia

What is the topic of this review? This article addresses the relationship between vagus nerve activity and malignant ventricular arrhythmias. It focuses on the clinical association of an impaired vagal tone in cardiac disease states with high mortality from sudden cardiac death and the potential underlying mechanisms. What advances does it highlight? The article summarizes the mounting evidence that vagal innervation in the cardiac ventricle plays a key direct role in the prevention of the initiation of ventricular fibrillation. Data are presented on the role that nitric oxide plays in mediating the effects of vagal protection against ventricular fibrillation, supporting the notion that a separate non-muscarinic, nitrergic population of vagal neurons is responsible for this protection. Sudden cardiac death remains a significant unresolved clinical problem, with many of the deaths being due to malignant ventricular arrhythmias. Markers of abnormal autonomic function have been shown to be strong prognostic predictors, highlighting the important relationship between reduced vagal tone and malignant ventricular arrhythmias, such as ventricular fibrillation, in cardiac patients. Exploring the mechanisms underlying the autonomic modulation of ventricular fibrillation, my group has shown that vagus nerve stimulation protects against ventricular fibrillation in the innervated isolated heart preparation. We have provided direct evidence that nitric oxide is released in the ventricle with cervical vagus nerve stimulation and NO mediates the antifibrillatory actions of vagus nerve stimulation in the ventricle. Classical physiology teaches that vagal postganglionic nerves modulate the heart via acetylcholine acting at muscarinic receptors and, dogmatically, that there is little vagal effect in the ventricle, as innervation was believed to be sparse. Mounting evidence from many species now supports the presence of a rich vagal innervation in the ventricle. Data from my group showing that the protective actions of vagus nerve stimulation against ventricular fibrillation and NO release are preserved in the presence of muscarinic block support the notion that a population of nitrergic neurons could be responsible. This potentially exploitable downstream pathway together with the availability of vagus nerve stimulators make it an exciting time to investigate the development of an effective strategy of vagal protection against ventricular fibrillation in the clinical setting.

Combining benefits of an adrenergic and a muscarinic blocker in a single formulation – A pharmacokinetic evaluation

A pharmacokinetic bioequivalence study was conducted in Asian subjects, to compare a fixed dose combination capsule single oral dose of alpha adrenoceptor blocker—Alfuzosin hydrochloride 10mg extended release and muscarinic antagonists—Solifenacin succinate 5mg against individually administered Xatral XL 10mg tablets (Alfuzosin) of Sanofi Synthelabo Limited, United Kingdom (UK) and Vesicare 5mg tablets (Solifenacin) of Astellas Pharma Limited, UK under fed conditions. Blood samples were collected pre-dose up to 72h post dose for determination of plasma Alfuzosin and Solifenacin concentrations and calculation of the pharmacokinetic parameters. ANOVA was performed on the log (natural)-transformed pharmacokinetic parameters. A 90% confidence interval for the ratios of the test and reference product averages (least square means) were calculated for alfuzosin and solifenacin. The 90% confidence intervals obtained for alfuzosin for Cmax, AUC0−t and AUC0−∞ were 102.74–122.75%, 95.84–116.96% and 95.82–116.76%, respectively. The 90% confidence intervals obtained for Solifenacin for Cmax, and AUC0−72 were 89.55–97.91% and 90.47–99.38%, respectively. Based on the results, the fixed dose combination was concluded to be bioequivalent to individually administered products.

Agonistic Autoantibodies as Vasodilators in Orthostatic Hypotension

Agonistic autoantibodies to the β-adrenergic and muscarinic receptors are a novel investigative and therapeutic target for certain orthostatic disorders. We have identified the presence of autoantibodies to β2-adrenergic and/or M3 muscarinic receptors by ELISA in 75% (15 of 20) of patients with significant orthostatic hypotension. Purified serum IgG from all 20 of the patients and 10 healthy control subjects were examined in a receptor-transfected cell-based cAMP assay for β2 receptor activation and β-arrestin assay for M3 receptor activation. There was a significant increase in IgG-induced activation of β2 and M3 receptors in the patient group compared with controls. A dose response was observed for both IgG activation of β2 and M3 receptors and inhibition of their activation with the nonselective β blocker propranolol and muscarinic blocker atropine. The antibody effects on β2 and/or M3 (via production of NO) receptor-mediated vasodilation were studied in a rat cremaster resistance arteriole assay. Infusion of IgG from patients with documented β2 and/or M3 receptor agonistic activity produced a dose-dependent vasodilation. Sequential addition of the β-blocker propranolol and the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester partially inhibited IgG-induced vasodilation (percentage of maximal dilatory response: from 57.7±10.4 to 35.3±4.6 and 24.3±5.8, respectively; P<0.01; n=3), indicating that antibody activation of vascular β2 and/or M3 receptors may contribute to systemic vasodilation. These data support the concept that circulating agonistic autoantibodies serve as vasodilators and may cause or exacerbate orthostatic hypotension.

Effects of acetylcholinesterase inhibitor paraoxon denote the possibility of non-quantal acetylcholine release in myocardium of different vertebrates

Effects of organophosphorous acetylcholinesterase inhibitor paraoxon were studied in the isolated atrial and ventricular myocardium preparations of a fish (cod), an amphibian (frog) and a mammal (rat) using the microelectrode technique. Incubation of isolated atrium with paraoxon (5×10(-6)-5×10(-5)M) caused significant reduction of action potential duration and marked slowing of sinus rhythm. These effects were abolished by muscarinic blocker atropine and therefore are caused by acetylcholine, which accumulates in the myocardium due to acetylcholinesterase inhibition even in the absence of vagal input. Hemicholinium III is a blocker of high affinity choline-uptake transporters, which are believed to mediate non-quantal release of acetylcholine from cholinergic terminals in different tissues. In the atrial myocardium of all the three studied species, hemicholinium III (10(-5)M) significantly suppressed all the effects of paraoxon. Blocker of parasympathetic ganglionic transmission hexamethonium bromide (10(-4)M) and inhibitor of vesicular acetylcholine transporters vesamicol (10(-5)M) failed to attenuate paraoxon effects. Among ventricular myocardium preparations of three species paraoxon provoked marked cholinergic effects only in frog, hemicholinium III abolished these effects effectively. We conclude that paraoxon stops degradation of acetylcholine in the myocardium and helps to reveal the effects of acetylcholine, which is continuously secreted from the cholinergic nerves in non-quantal manner. Thus, non-quantal release of acetylcholine in the heart is not specific only for mammals, but is also present in the hearts of different vertebrates.

The pharmacology of propionyl atropine methyl nitrate.

Abstract A series of esters of atropine and hyoscine, both in the tertiary and quaternary form, have been examined for their muscarinic and ganglionic blocking and neuromuscular transmission inhibiting activity. In addition the effect on the gastric secretion has been studied by the Shay rat method. Figures are also given for toxicities. In general, the pharmacology of the quaternary derivatives is qualitatively and quantitatively the same as the parent substances. The quaternary compounds retain a considerable amount of their tertiary activity but in addition become relatively strong ganglionic blocking agents. Most of the compounds reduce the volume and acidity of the gastric juice.

Some actions of 4,17a-dimethyl-4,17a-diaza-D-homo-5alpha-androstane dimethiodide (HS-342), a new neuromuscular blocking drug.

Abstract The effects of the newly-synthesized bisquaternary steroid 4,17a-dimethyl-4,17a-diaza-D-homo-5α-androstane dimethiodide (HS-342) have been investigated on the rat isolated phrenic nerve-hemidiaphragm, chick biventer cervicis and intact cat tibialis anterior muscle preparations, on the guinea-pig isolated ileum and on the rat isolated vas deferens preparations. The anticholinesterase activity of HS-342 was also measured. In the skeletal muscle preparations HS-342 exhibited properties of a post-junctionally active neuromuscular blocking agent of the non-depolarizing class. In the anaesthetized cat HS-342 exhibited approximately equal activity to that of tubocurarine, but its duration of action was about one-third of that of tubocurarine. HS-342 exhibited low anticholinesterase activity, low muscarinic blocking activity and apparently had no effect on the adrenergic neuroeffector junction studied.

Effect of muscarinic blocker on enhancing the action of fructus aurantii immaturus on intestinal myoelectric activity in dogs.

To investigate the effect of fructus aurantii immaturus (FAI) on small intestinal electrical activity in dogs. The effect of FAI was observed using a computerized electrophysiological method with the migrating myoelectric complex as a criterion. Fasted, healthy, and conscious dogs were given 100% FAI concentrated solution by gastrostogavage, and as soon as the effect on small intestinal electrical activity appeared, atropine was injected intramuscularly. The enhancing action of FAI was inhibited significantly by atropine, a cholinergic receptor antagonist. Both the number of spike bursts per cluster and the number of spikes per minute in phase II and III and the general cycle were decreased (P < 0.01), although the duration of phase II and the general cycle was prolonged. The effect of FAI might be related to the muscarinic receptors.

Cholinergic modulation of fast inhibitory and excitatory transmission to pedunculopontine thalamic projecting neurons.

The pedunculopontine nucleus (PPN) is part of the cholinergic arm of the reticular activating system, which is mostly active during waking and rapid-eye movement sleep. The PPN projects to the thalamus and receives cholinergic inputs from the laterodorsal tegmental nucleus and contralateral PPN. We employed retrograde labeling and whole cell recordings to determine the modulation of GABAergic, glycinergic, and glutamatergic transmission to PPN thalamic projecting neurons, and their postsynaptic responses to the nonspecific cholinergic agonist carbachol. M2 and M4 muscarinic receptor-modulated inhibitory postsynaptic responses were observed in 73% of PPN output neurons; in 12.9%, M1 and nicotinic receptor-mediated excitation was detected; and muscarinic and nicotinic-modulated fast inhibitory followed by slow excitatory biphasic responses were evident in 6.7% of cells. A significant increase in the frequency of spontaneous excitatory postsynaptic currents (EPSCs) and inhibitory postsynaptic currents during carbachol application was observed in 66.2% and 65.2% of efferent neurons, respectively. This effect was blocked by a M1 antagonist or nonselective muscarinic blocker, indicating that glutamatergic, GABAergic, and/or glycinergic neurons projecting to PPN output neurons are excited through muscarinic receptors. Decreases in the frequency of miniature EPSCs, and amplitude of electrical stimulation-evoked EPSCs, were blocked by a M2 antagonist, suggesting the presence of M2Rs at terminals of presynaptic glutamatergic neurons. Carbachol-induced multiple types of postsynaptic responses, enhancing both inhibitory and excitatory fast transmission to PPN thalamic projecting neurons through muscarinic receptors. These results provide possible implications for the generation of different frequency oscillations in PPN thalamic projecting neurons during distinct sleep-wake states.

Vascular Protection with Less Activation Evoked by Progressive Thermal Preconditioning in Adrenergic Receptor-Mediated Hypertension and Tachycardia

Thermal preconditioning may afford cardiovascular protection against oxidative injuries. However, hypertension and taychardia by sympathetic stimulation frequently occur during 420C whole body thermal preconditioning (TP). We aimed to develop a modified TP to achieve cardiovascular protection with to reduced cardiovascular stimulation in the rat. We used a progressive thermal preconditioning (PTP) with three-step 5-min immersion of male Wistar rats in 42 degrees C bath water. Treatment with phentolamine (alpha-adrenergic blocker), propranolol (beta-adrenergic blocker) and atropine (muscarinic cholinergic blocker) was used to evaluate the effect and mechanism of PTP on systemic hemodynamics. Protective function was evaluated by FeCl3-induced acute femoral arterial occlusion (TTO) and heat shock protein 70 expression. Our results show that TP enhanced body temperature, hypertension and tachycardia. However, PTP produced a similar increase in body temperature with significantly less enhancement of hypertension and tachycardia when compared with the TP group. TP- or PTP-induced increase of blood pressure and heart rate was inhibited by phentolamine and propranolol, respectively. PTP-induced attenuation of changes in hemodynamic parameters was via alpha- and beta-adrenergic inhibition. FeCl3 induced femoral arterial injury indicated by TTO at 416 +/- 51 sec in the control rats. After 24 h of TP or PTP treatment with or without adrenergic blocker treatment, TP or PTP upregulated similar femoral arterial heat shock protein 70 expression and significantly (P < 0.05) delayed FeCl3-induced femoral TTO to a similar degree. PTP may provide vascular protection against oxidative injuries with less activation in alpha-adrenergic receptor-mediated hypertension and beta-adrenergic receptor-mediated tachycardia.

Pouch colon associated with anorectal malformations fails to show spontaneous contractions but responds to acetylcholine and histamine in vitro

Purpose Congenital pouch colon (CPC) associated with anorectal malformation (ARM) is most commonly reported from Northern India. So far, no physiologic study comparing the detailed contractile status of CPC with non-CPC conditions are available. The present article deals with the contractile study and histopathologic observations in CPC, which may be useful for better surgical management. Methods Freshly excised 12 neonatal CPC and similar number of non-CPC (control) specimens were transferred to ice-cold (4°C-6°C) Krebs-Ringer solution bubbled with 100% oxygen. Longitudinally prepared 2 to 4 colonic strips were obtained from central part of each specimen and subjected to the contraction recording after exposure to cumulative concentrations of acetylcholine (ACh) and histamine. Acetylcholine-induced contractions were evaluated after application of atropine (muscarinic blocker), and histaminergic contractions were recorded after pheniramine (H 1 blocker), lignocaine (neuronal blocker), and atropine. Histopathologic observations were made by using H&E and Masson trichrome stains. Results Control specimens showed spontaneous contractions, but CPC strips did not. Both control and CPC responded to ACh and histamine. The response to histamine was greater ( P < .05) in CPC as compared to control, whereas the response to ACh was more ( P < .05) in control. In CPC, response of histamine (100 μmol/L) was blocked by pheniramine (0.32 mmol/L) and lignocaine (4 mmol/L) by 97% and 80%, respectively, and enhanced by 57% after preapplication of atropine (10 μmol/L). Acetylcholine (100 μmol/L)-induced contractions were attenuated (86%) in presence of atropine. Histopathologic examination showed fewer mature ganglion cells with various changes in muscle layers including fibrosis, disruption, hypertrophy, atrophy, and constriction bands. Conclusion Congenital pouch colon associated with ARM lacks normal spontaneous contractions but retains ACh and histamine-induced contractility. In view of the functional and histologic abnormalities, we propose that CPC associated with ARM is an abnormally functional and developed tissue. Therefore, resection of the pouch should be considered for better functional outcome of the remaining bowel.