Use of chlorisondamine to assess the neurogenic contribution to blood pressure in mice: An evaluation of method.

Chlorisondamine (CSD) has been used to assess the neurogenic contribution to blood pressure (BP) and vasomotor sympathetic tone in animal models. It is assumed that the reduction in BP following CSD administration is associated to decreases in cardiac output (CO) and peripheral resistance, reflecting cardiac and vasomotor sympathetic tone, respectively. Surprisingly, this has not been characterized experimentally in mice, despite the extensive use of this animal model in cardiovascular research. We hypothesize that a specific dose of CSD can selectively block the sympathetic vasomotor tone. To test this hypothesis, we evaluated the effects of different doses of CSD (intraperitoneal) on BP and heart rate (HR) using telemetry, and on CO using echocardiography. BP and HR in normotensive C57Bl/6J mice reduced to a similar extent by all CSD doses tested (1-6 mg/kg). CSD at 6 mg/kg also reduced CO without affecting left ventricular stroke volume or fractional shortening. On the other hand, lower doses of CSD (1 and 2 mg/kg) produced significantly larger BP and HR reductions in DOCA-salt-induced hypertensive mice, indicating a greater neurogenic BP response. In addition, all doses of CSD reduced CO in hypertensive mice. Our data suggest that the BP response to CSD in mice likely reflects reduced CO and vasomotor sympathetic tone. We conclude that CSD can be used to assess the neurogenic contribution to BP in mice but may not be appropriate for specifically estimating vasomotor sympathetic tone.

Soluble Prorenin Receptor Increases Blood Pressure in High Fat-Fed Male Mice.

Obesity-related hypertension is a major public health concern. We recently demonstrated that plasma levels of the soluble form of the prorenin receptor (sPRR) were elevated in obesity-associated hypertension. Therefore, in the present study, we investigated the contribution of sPRR to blood pressure (BP) elevation in the context of obesity. High fat-fed C57BL/6 male mice were infused with vehicle or sPRR (30 µg/kg per day) via subcutaneously implanted osmotic minipump for 4 weeks. BP parameters were recorded using radiotelemetry devices. Male mice infused with sPRR exhibited higher systolic BP and mean arterial pressure and lower spontaneous baroreflex sensitivity than mice infused with vehicle. To define mechanisms involved in systolic BP elevation, mice were injected with an AT1R (Ang II [angiotensin II] type 1 receptor) antagonist (losartan), a muscarinic receptor antagonist (atropine), a ß-adrenergic antagonist (propranolol), and a ganglionic blocker (chlorisondamine). Losartan did not blunt sPRR-induced elevation in systolic BP. Chlorisondamine treatment exacerbated the decrease in mean arterial pressure in male mice infused with sPRR. These results demonstrated that sPRR induced autonomic nervous dysfunction. Interestingly, plasma leptin levels were increased in high fat-fed C57BL/6 male mice infused with sPRR. Overall, our results indicated that sPRR increased systolic BP through an impairment of the baroreflex sensitivity and an increase in the sympathetic tone potentially mediated by leptin in high fat-fed C57BL/6 male mice.

Species specificity of rat and human α7 nicotinic acetylcholine receptors towards different classes of peptide and protein antagonists.

Peptide and protein neurotoxins, such as α-conotoxins from Cone snails and α-neurotoxins from snake venoms, are excellent tools to identify distinct nicotinic acetylcholine receptor (nAChR) subtypes. Here we compared the rat/human species specificity of α7 nAChR towards peptide and protein neurotoxins and found that α-conotoxin analogues [K11A]TxIB and [H5D]RegIIA are much more potent on the rat versus human α7 receptor expressed in Xenopus oocytes. In the hope to determine the key residue responsible for the difference in α-conotoxin analogues affinities, ten single mutants of rat α7 nAChR were obtained because there are 10 differences in the extracellular ligand-binding domains of these species, and only K185R mutation decreased the affinity for α-conotoxins [K11A]TxIB and [H5D]RegIIA, down to their low affinities for human α7 nAChR. On the other hand, the reverse mutation R185K in human α7 nAChR resulted in the greatest increase in the affinity for both conotoxins, while a double mutation hα7[S183N, R185K] made the potency of the receptor for them as high as that of rat α7 nAChR. The effects of mutations at position 185 were investigated also with some other α-conotoxins and cobra venom α-cobratoxin and found to have similar but much less pronounced effects on their species specificity. Molecular modeling provided possible explanation for the high species selectivity of [K11A]TxIB and [H5D]RegIIA towards α7 nAChR, opening the new way for design of their analogues with improved affinity to the human receptor.

Nicotine versus 6-hydroxy-l-nicotine against chlorisondamine induced memory impairment and oxidative stress in the rat hippocampus.

6-Hydroxy-l-nicotine (6HLN), a nicotine derivative from nicotine degradation by Arthrobacter nicotinovorans pAO1 strain was found to improve behavioral deficits and to reverse oxidative stress in the rat hippocampus. Rats were given CHL (10mg/kg, i.p.) were used as an Alzheimer's disease-like model. The nicotine (0.3mg/kg) and 6HLN (0.3mg/kg) were administered alone or in combination in the CHL-treated rats. Memory-related behaviors were evaluated using Y-maze and radial arm-maze tests. The antioxidant enzymes activity and the levels of the biomarkers of oxidative stress were measured in the hippocampus. Statistical analyses were performed using two-way ANOVA and Tukey's post hoc test. F values for which p<0.05 were regarded as statistically significant. CHL-caused memory deficits and oxidative stress enhancing were observed. Both nicotine and 6HLN administration attenuated the cognitive deficits and recovered the antioxidant capacity in the rat hippocampus of the CHL rat model. Our results suggest that 6HLN versus nicotine confers anti-amnesic properties in the CHL-induced a rat model of memory impairment via reversing cholinergic function and decreasing brain oxidative stress, suggesting the use of this compound as an alternative agent in AD treatment.

Role of α7-Nicotinic Acetylcholine Receptors of B Cells in the Immunotoxic Effect of Organophosphorus Compounds.

Experiments on white non-inbred rats demonstrated that treatment with organophosphorus compound dimethyl dichlorovinyl phosphate (DDVP) decreased T cell-independent antibody production by B cells and blood levels of IL-10 and IL-12; a similar effect was produced by GTS-21, a selective agonist of α7-nicotinic acetylcholine receptor. N-nicotinic receptor antagonist chlorisondamine in combination with DDVP partially prevented suppression of antibody production in comparison with the effect observed during intoxication with DDVP.

Central Insulin Action Activates Kupffer Cells by Suppressing Hepatic Vagal Activation via the Nicotinic Alpha 7 Acetylcholine Receptor.

Central insulin action activates hepatic IL-6/STAT3 signaling, which suppresses the gene expression of hepatic gluconeogenic enzymes. The vagus nerve plays an important role in this centrally mediated hepatic response; however, the precise mechanism underlying this brain-liver interaction is unclear. Here, we present our findings that the vagus nerve suppresses hepatic IL-6/STAT3 signaling via α7-nicotinic acetylcholine receptors (α7-nAchR) on Kupffer cells, and that central insulin action activates hepatic IL-6/STAT3 signaling by suppressing vagal activity. Indeed, central insulin-mediated hepatic IL-6/STAT3 activation and gluconeogenic gene suppression were impeded in mice with hepatic vagotomy, pharmacological cholinergic blockade, or α7-nAchR deficiency. In high-fat diet-induced obese and insulin-resistant mice, control of the vagus nerve by central insulin action was disturbed, inducing a persistent increase of inflammatory cytokines. These findings suggest that dysregulation of the α7-nAchR-mediated control of Kupffer cells by central insulin action may affect the pathogenesis of chronic hepatic inflammation in obesity.

Gintonin facilitates catecholamine secretion from the perfused adrenal medulla

The present study was designed to investigate the characteristics of gintonin, one of components isolated from Korean Ginseng on secretion of catecholamines (CA) from the isolated perfused model of rat adrenal gland and to clarify its mechanism of action. Gintonin (1 to 30 µg/ml), perfused into an adrenal vein, markedly increased the CA secretion from the perfused rat adrenal medulla in a dose-dependent fashion. The gintonin-evoked CA secretion was greatly inhibited in the presence of chlorisondamine (1 µM, an autonomic ganglionic bloker), pirenzepine (2 µM, a muscarinic M₁ receptor antagonist), Ki14625 (10 µM, an LPA₁/₃ receptor antagonist), amiloride (1 mM, an inhibitor of Na⁺/Ca²⁺ exchanger), a nicardipine (1 µM, a voltage-dependent Ca²⁺ channel blocker), TMB-8 (1 µM, an intracellular Ca²⁺ antagonist), and perfusion of Ca²⁺-free Krebs solution with 5mM EGTA (a Ca²⁺chelater), while was not affected by sodium nitroprusside (100 µM, a nitrosovasodialtor). Interestingly, LPA (0.3~3 µM, an LPA receptor agonist) also dose-dependently enhanced the CA secretion from the adrenal medulla, but this facilitatory effect of LPA was greatly inhibited in the presence of Ki 14625 (10 µM). Moreover, acetylcholine (AC)-evoked CA secretion was greatly potentiated during the perfusion of gintonin (3 µg/ml). Taken together, these results demonstrate the first evidence that gintonin increases the CA secretion from the perfused rat adrenal medulla in a dose-dependent fashion. This facilitatory effect of gintonin seems to be associated with activation of LPA- and cholinergic-receptors, which are relevant to the cytoplasmic Ca²⁺ increase by stimulation of the Ca²⁺ influx as well as by the inhibition of Ca²⁺ uptake into the cytoplasmic Ca²⁺ stores, without the increased nitric oxide (NO). Based on these results, it is thought that gintonin, one of ginseng components, can elevate the CA secretion from adrenal medulla by regulating the Ca²⁺ mobilization for exocytosis, suggesting facilitation of cardiovascular system. Also, these findings show that gintonin might be at least one of ginseng-induced hypertensive components.

Cerebrospinal Fluid Hypernatremia Elevates Sympathetic Nerve Activity and Blood Pressure via the Rostral Ventrolateral Medulla.

Elevated NaCl concentrations of the cerebrospinal fluid increase sympathetic nerve activity (SNA) in salt-sensitive hypertension. Neurons of the rostral ventrolateral medulla (RVLM) play a pivotal role in the regulation of SNA and receive mono- or polysynaptic inputs from several hypothalamic structures responsive to hypernatremia. Therefore, the present study investigated the contribution of RVLM neurons to the SNA and pressor response to cerebrospinal fluid hypernatremia. Lateral ventricle infusion of 0.15 mol/L, 0.6 mol/L, and 1.0 mol/L NaCl (5 µL/10 minutes) produced concentration-dependent increases in lumbar SNA, adrenal SNA, and arterial blood pressure, despite no change in splanchnic SNA and a decrease in renal SNA. Ganglionic blockade with chlorisondamine or acute lesion of the lamina terminalis blocked or significantly attenuated these responses, respectively. RVLM microinjection of the gamma-aminobutyric acid (GABAA) agonist muscimol abolished the sympathoexcitatory response to intracerebroventricular infusion of 1 mol/L NaCl. Furthermore, blockade of ionotropic glutamate, but not angiotensin II type 1, receptors significantly attenuated the increase in lumbar SNA, adrenal SNA, and arterial blood pressure. Finally, single-unit recordings of spinally projecting RVLM neurons revealed 3 distinct populations based on discharge responses to intracerebroventricular infusion of 1 mol/L NaCl: type I excited (46%; 11/24), type II inhibited (37%; 9/24), and type III no change (17%; 4/24). All neurons with slow conduction velocities were type I cells. Collectively, these findings suggest that acute increases in cerebrospinal fluid NaCl concentrations selectively activate a discrete population of RVLM neurons through glutamate receptor activation to increase SNA and arterial blood pressure.

Chlorisondamine, a sympathetic ganglionic blocker, moderates the effects of whole-body irradiation (WBI) on early host defense to a live bacterial challenge.

There is a growing consensus that long-term deficits in the brain are due to dynamic interactions between multiple neural and immune cell types. Specifically, radiation induces an inflammatory response, including changes in neuromodulatory pro- and anti-inflammatory cytokine secretion. The purpose of this study was to establish that there is sympathetic involvement in radiation-induced decrements early in in vivo immune function host defense. Female, 8-9 week-old C57BL/6J mice were exposed to whole-body irradiation (WBI). There were 8 groups with radiation (0 vs. 3 Gy protons), immune challenge (Escherichia coli) and exposure to the sympathetic ganglionic blocker, chlorisondamine (1 mg/kg weight, i.p.), as independent variables. Ten days post-irradiation, mice were inoculated with E. coli intraperitoneally and sacrificed 90-120 min later. The data suggest that radiation-induced changes in immune function may in part be mediated by the sympathetic nervous system. Briefly, we found that radiation augments the bacteria-induced inflammatory cytokine response, particularly those cytokines involved in innate immunity. However, this augmentation can be reduced by the ganglionic blockade.

Melanocortin-4 receptor agonists alleviate intestinal dysfunction in secondary intra-abdominal hypertension rat model.

BACKGROUND: Intra-abdominal hypertension (IAH) is a potentially life-threatening disease. Melanocortin-4 (MC4) receptor activation exhibits life-saving properties. The aim of the present study was to examine whether treatment with the MC4 receptor agonist RO27-3225 ameliorates intestinal injury in IAH rats. METHODS: A total of 72 male Sprague-Dawley rats were randomized into six groups. Group 1 was the sham group. Group 2, the sham + RO group, received RO27-3225 (180 µg/kg, intraperitoneally). IAH was induced in group 3, the IAH group, by blood draw (mean arterial pressure = 30 mm Hg for 90 min) followed by shed blood and/or Ringer solution reinfusion. Intra-abdominal pressure was increased to 20 mm Hg by injecting air into the peritoneal cavity. Group 4, the RO group, was administered RO27-3225 at 5 min after blood draw. Groups 5 and 6 were the chlorisondamine (Chl) and HS024 groups, in which the rats were pretreated with the nicotinic acetylcholine receptor antagonist Chl or selective MC4 receptor antagonist (HS024), respectively, at 2 min before RO27-3225 was administered. RESULTS: RO27-3225 restored mean arterial pressure, reduced tumor necrosis factor-α, and interleukin-1ß messenger RNA expression increased by IAH, alleviated histologic damage, and improved superoxide dismutase activity in the intestine. Compared with the IAH group, the levels of intestinal fatty acid-binding protein, intestinal edema and intestinal permeability were lower in the RO group. Furthermore, the RO27-3225 treatment increased the expression of Rho-associated coiled-coil-containing protein kinase 1 and phosphorylated myosin light chain. Chl and HS024 abrogated the protective effects of RO27-3225. CONCLUSIONS: These data indicate that the MC4 receptor agonist counteracts the intestinal inflammatory response, ameliorating intestinal injury in experimental secondary IAH by MC4 receptor-triggered activation of the cholinergic anti-inflammatory pathway. It may represent a promising strategy for the treatment of IAH in the future.

Prevention of hemolysis-induced organ damage by nutritional activation of the vagal anti-inflammatory reflex*.

OBJECTIVES: Acute hemolysis is associated with organ damage, inflammation, and impaired vascular function. Stimulation of the cholecystokinin-1 receptor-dependent vagal anti-inflammatory reflex with lipid-rich enteral nutrition was demonstrated to prevent tissue damage and attenuate inflammation. This study investigates the effects of nutritional activation of the vagal anti-inflammatory reflex on organ integrity, systemic inflammation, and microcirculation during hemolysis. DESIGN: Prospective randomized controlled study. SETTING: University research unit. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: Intravascular hemolysis was simulated by infusion of prelysed erythrocytes. Animals were fasted or received lipid-rich enteral nutrition. Pegylated (PEG)-CCK9A, A70104 (a cholecystokinin-1 receptor antagonist), and chlorisondamine (a nicotinic acetylcholine receptor antagonist) were applied to investigate involvement of the vagal reflex. MEASUREMENTS AND MAIN RESULTS: Nutritional intervention reduced hemolysis-related renal tubular cell damage, hepatocyte damage, ileal leakage of horseradish peroxidase, and bacterial translocation compared with food deprivation (all p < 0.05). Also circulating interleukin (IL)-6 levels were decreased by enteral nutrition (p < 0.05). Blockage of the cholecystokinin-1 receptor or the nicotinic acetylcholine receptor reversed the protective nutritional effects compared with vehicle (p < 0.05), whereas PEG-CCK9 mimicked the impact of enteral feeding in fasted animals (p < 0.05). Furthermore, nutritional intervention increased renal, hepatic, and intestinal blood flow compared with fasting (all p < 0.05), as evaluated using fluorescent microspheres. CONCLUSIONS: Nutritional activation of the vagal anti-inflammatory reflex preserves tissue integrity and attenuates systemic inflammation in a rodent model of acute hemolysis. In addition, lipid-rich nutrition improves renal, hepatic, and intestinal microcirculation. These findings implicate stimulation of the autonomic nervous system by nutritional means as a potential therapy to prevent complications of acute hemolysis. (Crit Care Med 2013; 41:e361-e367).

Neural control of arterial pressure variability in the neuromuscularly blocked rat.

The baroreflexes stabilize moment-to-moment arterial pressure. Sinoaortic denervation (SAD) of the baroreflexes results in a large increase in arterial pressure variability (APV) across various species. Due to an incomplete understanding of the nonlinear interactions between central and peripheral systems, the major source of APV remains controversial. While some studies suggested that the variability is endogenous to the central nervous system (CNS), others argued that peripheral influences may be the main source. For decades, abnormal cardiovascular variability has been associated with a number of cardiovascular diseases including hypertension, heart failure, and stroke. Delineating mechanisms of the APV is critical for the improvement of current strategies that use APV as a clinical tool for the diagnosis and prognosis of cardiovascular diseases. In this study, with a unique chronic neuromuscularly blocked (NMB) rat preparation that largely constrains peripheral influences, we determined the CNS contribution to the post-SAD APV. First, we confirmed that SAD significantly increased APV in the NMB rat, then demonstrated that post-SAD ganglionic blockade substantially reduced APV, and subsequent intravenous infusions of phenylephrine and epinephrine (in presence of ganglionic blockade) only slightly increased APV. These data suggest that the CNS is an important source, and skeletal activity, thermal challenges or other forms of peripherally generated cardiovascular stress are not required for the post-SAD APV. In addition, we showed that bilateral aortic denervation produced a larger increase in APV than bilateral carotid sinus denervation, suggesting that the aortic baroreflex plays a more dominant role in the control of APV than the carotid sinus.

Drug metabolism within the brain changes drug response: selective manipulation of brain CYP2B alters propofol effects.

Drug-metabolizing cytochrome P450 (CYPs) enzymes are expressed in the liver, as well as in extrahepatic tissues such as the brain. Here we show for the first time that drug metabolism by a CYP within the brain, illustrated using CYP2B and the anesthetic propofol (2, 6-diisopropylphenol, Diprivan), can meaningfully alter the pharmacological response to a CNS acting drug. CYP2B is expressed in the brains of animals and humans, and this CYP isoform is able to metabolize centrally acting substrates such as propofol, ecstasy, and serotonin. Rats were given intracerebroventricularly (i.c.v.) injections of vehicle, C8-xanthate, or 8-methoxypsoralen (CYP2B mechanism-based inhibitors) and then tested for sleep time following propofol (80 mg/kg intraperitoneally). Both inhibitors significantly increased sleep-time (1.8- to 2-fold) and brain propofol levels, while having no effect on plasma propofol levels. Seven days of nicotine treatment can induce the expression of brain, but not hepatic, CYP2B, and this induction reduced propofol sleep times by 2.5-fold. This reduction was reversed in a dose-dependent manner by i.c.v. injections of inhibitor. Sleep times correlated with brain (r=0.76, P=0.0009), but not plasma (r=0.24, P=0.39) propofol concentrations. Inhibitor treatments increased brain, but not plasma, propofol levels, and had no effect on hepatic enzyme activity. These data indicate that brain CYP2B can metabolize neuroactive substrates (eg, propofol) and can alter their pharmacological response. This has wider implications for localized CYP-mediated metabolism of drugs, neurotransmitters, and neurotoxins within the brain by this highly variable enzyme family and other CYP subfamilies expressed in the brain.

Rat brain CYP2B induction by nicotine is persistent and does not involve nicotinic acetylcholine receptors.

CYP2B is a drug-metabolizing enzyme expressed in the liver and brain that metabolizes a variety of centrally acting drugs (e.g. propofol, bupropion and nicotine), endogenous neurochemicals (e.g. serotonin and testosterone) and toxins (e.g. chlorpyrifos). Human CYP2B6 is found at higher levels in the brains of smokers, and 7-day nicotine treatment induces rat brain CYP2B while not altering hepatic CYP2B. We characterized the time course of rat brain CYP2B induction by nicotine and determined if nicotinic acetylcholine receptors (nAChRs) mediated this induction. Rats were treated once daily with 1mg/kg nicotine base or saline s.c. for 1 or 7days and sacrificed from 30minutes to 7days after the last injection. One-day nicotine treatment did not induce brain CYP2B, whereas 7-day nicotine treatment significantly increased CYP2B expression for up to 24hours in the frontal cortex and brainstem; these levels returned to baseline by 7days post-treatment. CYP2B activity was also higher at 24hours in these regions. No change was seen in the cerebellar CYP2B levels or in vivo activity following nicotine treatment. Time of day of treatment and sacrifice altered the magnitude of brain CYP2B induction while chlorisondamine, an irreversible nAChR blocker, pre-treatment did not block CYP2B induction. Seven-day nicotine treatment resulted in an induction of rat brain CYP2B protein and in vivo activity for up to 24hours, which would suggest greater local drug metabolism by brain CYP2B. Humans or animals exposed to nicotine may have altered therapeutic drug response, brain levels of neurotransmitters and/or neurotoxicity.

Lipid-enriched enteral nutrition controls the inflammatory response in murine Gram-negative sepsis.

OBJECTIVES: Controlling the inflammatory cascade during sepsis remains a major clinical challenge. Recently, it has become evident that the autonomic nervous system reduces inflammation through the vagus nerve. The current study investigates whether nutritional stimulation of the autonomic nervous system effectively attenuates the inflammatory response in murine Gram-negative sepsis. DESIGN: Controlled in vivo and ex vivo experimental study. SETTINGS: Research laboratory of a university hospital. SUBJECTS: Male C57bl6 mice. INTERVENTIONS: Mice were intraperitoneally challenged with lipopolysaccharide derived from Escherichia coli. Before lipopolysaccharide administration, mice were fasted or enterally fed either lipid-rich nutrition or low-lipid nutrition. Antagonists to cholecystokinin receptors or nicotinic receptors were administered before lipopolysaccharide administration. Blood and tissue samples were collected at 90 mins. Mesenteric afferent discharge was determined in ex vivo preparations in response to both nutritional compositions. MEASUREMENTS AND MAIN RESULTS: Both lipid-rich and low-lipid nutrition dose-dependently reduced lipopolysaccharide-induced tumor necrosis factor-α release (high dose: both 1.4 ± 0.4 ng/mL) compared with fasted mice (3.7 ± 0.8 ng/mL; p < .01). The anti-inflammatory effect of both nutritional compositions was mediated through cholecystokinin receptors (p < .01), activation of mesenteric vagal afferents (p < .05), and peripheral nicotinic receptors (p < .05). Lipid-rich nutrition attenuated the inflammatory response at lower dosages than low-lipid nutrition, indicating that enrichment of enteral nutrition with lipid augments the anti-inflammatory potential. Administration of lipid-rich nutrition prevented endotoxin-induced small intestinal epithelium damage and reduced inflammation in the liver and spleen compared with fasted (all p < .01) and low-lipid nutrition controls (all p < .05). CONCLUSIONS: The current study demonstrates that lipid-rich nutrition attenuates intestinal damage and systemic as well as organ-specific inflammation in murine Gram-negative sepsis through the nutritional vagal anti-inflammatory pathway. These findings implicate enteral administration of lipid-enriched nutrition as a promising intervention to modulate the inflammatory response during septic conditions.

Increase in locomotor activity after acute administration of the nicotinic receptor agonist 3-bromocytisine in rats.

Nicotinic acetylcholine receptors influence striatal dopaminergic activity and its outcome on motor behavior. For these reasons, nicotinic receptors have been considered as therapeutically relevant targets for Parkinson's disease, in which a dramatic loss of dopamine affects motor functions. The aim of the present work was to compare the effects on locomotor activity induced by the nicotinic agonist cytisine and two brominated derivatives, 5- and 3-bromocytisine (5-BrCy and 3-BrCy) using nicotine for comparison. After acute systemic administration of the agonists only 3-BrCy induced an increase in locomotor activity. To study the mechanism of action involved in this increase we co-administered 3-BrCy with the nicotinic antagonist mecamylamine and also examined 3-BrCy's effects in rats pre-treated with the long acting nicotinic antagonist chlorisondamine, administered directly in the dorsal and ventral striatum. We studied the role of the dopaminergic system by co-administration of the D2 dopamine receptor antagonist, haloperidol. The results indicate that the increase in motor activity elicited by 3-BrCy was mediated by nicotinic receptors in the dorsal and ventral striatum and depends on the interaction of nicotinic receptors with the dopaminergic system. We conclude that 3-BrCy might be a new tool to study the modulation of the dopaminergic system by nicotinic receptors and their behavioral implications.

Prenatal high salt programs enhanced sympathoadrenal activation of the cardiovascular response to restraint.

We recently reported that feeding Sprague Dawley rats a high-salt diet during pregnancy programmed an exaggerated pressor and tachycardic response to restraint in adult female offspring. In the present investigation, a pharmacologic approach was used to determine the contribution of the sympathoadrenal system to the exaggerated response. Injection of a cocktail containing a ganglionic blocker (chlorisondamine) and a beta-adrenoceptor antagonist (propranolol) prevented the stress-induced tachycardia and increase in blood pressure and abolished the difference between high-salt and normal-salt offspring. These data suggest that the prenatal high salt programmed a sympathoadrenal hyperresponsiveness to restraint stress.

Morphine and ABT-594 (a nicotinic acetylcholine agonist) exert centrally mediated antinociception in the rat cyclophosphamide cystitis model of visceral pain.

UNLABELLED: A visceral pain model incorporating use of cyclophosphamide (CP) to induce bladder inflammation has been described. CP treatment in rats produces changes in behavior (abnormal postures and eye closure) and respiration rate indicative of visceral pain. We characterized the dose-dependency and progression of CP-induced cystitis pain after intraperitoneal (i.p.) CP. The behavioral and respiration rate changes were ameliorated by systemic morphine and ABT-594 [(R)-5-(2-azetidinylmethoxy)-2-chloropyridine], a neuronal nicotinic acetylcholine receptor agonist, in a manner reversible by naloxone and mecamylamine, respectively. Sites of antinociceptive actions of morphine and ABT-594 were investigated using systemic, intrathecal (i.t.), or intracerebroventricular (i.c.v.) administration of blood-brain barrier impenetrant antagonists. Naloxone methiodide produced a complete antagonism of morphine antinociception after i.c.v. but not i.p. or i.t. administration. Chlorisondamine blocked ABT-594 antinociception after i.c.v. but not i.p. administration. Further pharmacological characterization of behavioral and respiration changes in CP-cystitis was performed using standard analgesics. The alpha(2)-adrenoceptor agonist clonidine produced a weak attenuation of CP-pain behavior. NSAIDs (ibuprofen, acetaminophen, and celecoxib) and anticonvulsants (gabapentin and lamotrigine) were without effect. These results demonstrate that morphine and ABT-594 produce antinociception in CP-cystitis by a predominantly supraspinal site of action, and that mechanisms producing robust centrally-mediated antinociception could be beneficial in cystitis pain. PERSPECTIVE: In this article, potential antinociceptive effects of a variety of pharmacological agents were evaluated in a rat cystitis pain model. Morphine and a nicotinic acetylcholine receptor agonist ABT-594 were found to exert potent antinociception in this model. Findings presented here aid identification of agents to treat cystitis pain in the clinic.

Effects of dipeptidyl peptidase iv inhibition on arterial blood pressure.

1. The aim of the present study was to determine whether inhibition of dipeptidyl peptidase IV (DPP IV) elevates arterial blood pressure and whether any such effect is dependent on genetic background, the sympathetic nervous system and Y(1) receptors. The rationale behind this study was that: (i) neuropeptide (NP) Y(1-36) and peptide YY(1-36) (PYY(1-36)) are endogenous Y(1) receptor agonists and are metabolised by DPP IV to NPY(3-36) and PYY(3-36), which are not Y(1) but rather selective Y(2) receptor agonists; (ii) Y(1) receptors mediate vasoconstriction, whereas Y(2) receptors have little effect on vascular tone; (iii) vaso-constrictor effect of the Y(1) receptor is enhanced in spontaneously hypertensive rats (SHR) compared with normotensive Wistar-Kyoto (WKY) rats; and (iv) NPY(1-36) is released from sympathetic nerve terminals. 2. We examined the effects of acute administration of 3-N-[(2S,3S)-2-amino-3-methylpentanoyl]-1,3-thiazolidine (P32/98; a DPP IV inhibitor) on arterial blood pressure in anaesthetized adult SHR and WKY rats in the absence and presence of either captopril, hydralazine or chlorisondamine to lower basal mean arterial blood pressure (MABP) by different mechanisms (inhibition of angiotensin-converting enzyme, direct vasodilation and ganglionic blockade, respectively). 3. In naïve SHR with severely elevated basal blood pressures (MABP = 176 +/- 3 mmHg; n = 4), i.v. boluses (1, 3 and 10 mg/kg) of P32/98 did not affect blood pressure. 4. When basal blood pressure was reduced by pretreatment of SHR with either captopril (30 mg/kg, i.v.; MABP = 116 +/- 3 mmHg; n = 9) or hydralazine (5 mg/kg, i.p.; MABP = 84 +/- 3 mmHg; n = 7), P32/98 (1, 3 and 10 mg/kg) caused significant dose-related increases in arterial blood pressure (4 +/- 2, 10 +/- 2 and 12 +/- 3 mmHg in the captopril-pretreated group, respectively (P < 0.01); 5 +/- 2, 8 +/- 3 and 11 +/- 4 mmHg in the hydralazine-pretreated group, respectively (P < 0.01)). 5. The increases in arterial blood pressure induced by P32/98 in captopril- or hydralazine-pretreated SHR were entirely blocked by pretreatment with the selective Y(1) receptor antagonist N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-d-arginine amide (BIBP 3226; 6 mg/kg per h). 6. When basal blood pressure was reduced in SHR by pretreatment with chlorisondamine (10 mg/kg, s.c.; MABP = 108 +/- 4 mmHg; n = 7), inhibition of DPP IV with P32/98 did not affect arterial blood pressure. Basal heart rate in chlorisondamine-treated SHR was significantly reduced compared with naïve SHR, captopril-pretreated SHR and hydralazine-pretreated SHR, indicating effectiveness of ganglionic blockade. 7. Unlike the results in genetically hypertensive animals, in normotensive WKY rats pretreated with captopril (30 mg/kg, i.v.; MABP = 81 +/- 4 mmHg; n = 6), or hydralazine (5 mg/kg, i.p.; MABP = 63 +/- 4 mmHg; n = 4) or chlorisondamine (10 mg/kg, s.c.; MABP = 63 +/- 4 mmHg; n = 5), P32/98 did not affect arterial blood pressure. 8. We conclude that, in genetically susceptible animals, inhibition of DPP IV increases arterial blood pressure via Y(1) receptors when elevated blood pressure is reduced with antihypertensive drugs provided that the sympathetic nervous system is functional. The results suggest vigilance because DPP IV inhibitors are used more widely in hypertensive patients treated with antihypertensive drugs.

The effect of the cholinergic anti-inflammatory pathway on experimental colitis.

Inflammatory bowel diseases (IBD) are characterized by proinflammatory cytokines, tissue damage and loss of neuron in inflamed mucosa, which implies the cholinergic anti-inflammatory pathway may be destroyed during the process of inflammatory response. In the study, we identified the effect of cholinergic agonist as anabaseine (AN) and nicotinic receptor antagonist as chlorisondamine diiodide (CHD) on trinitrobenzene sulfonic acid (TNBS)-induced colitis, to investigate the potential therapeutic effect of the cholinergic anti-inflammatory pathway on IBD. Experimental colitis was induced by TNBS at day 1, 10 mug AN or 1.5 mug CHD was injected i.p. to mouse right after the induction of colitis, and repeated on interval day till the mice were sacrificed at day 8. Colonic inflammation was examined by histological analysis, myeloperoxidase (MPO) activity, and the production of tumour necrosis factor (TNF)-alpha in tissue. Lamina propria mononuclear cells (LPMC) were isolated, and NF-kappaB activation was detected by western blot. The mice with colitis treated by AN showed less tissue damage, less MPO activity, less TNF-alpha production in colon, and inhibited NF-kappaB activation in LPMC, compared with those mice with colitis untreated, whereas the mice with colitis treated by CHD showed the worst tissue damage, the highest MPO activity, the highest TNF-alpha level, and enlarged NF-kappaB activation in LPMC. Agonist of the cholinergic anti-inflammatory pathway inhibits colonic inflammatory response by downregulating the production of TNF-alpha, and inhibiting NF-kappaB activation, which suggests that modulating the cholinergic anti-inflammatory pathway may be a new potential management for IBD.