Antinociception with intrathecal alpha-methyl-5-hydroxytryptamine,a 5-hydroxytryptamine 2A/2C receptor agonist,in two rat models of sustained pain

Type 2 serotonin (5-hydroxytryptamine [5-HT](2)) receptors in the spinal cord have been reported to mediate antinociception using pain threshold tests, but little is known about the actions of spinal 5-HT(2) receptors in sustained pain. In rats, we examined antinociceptive effects of the intrathecal administration of a 5-HT(2A/2C) receptor agonist, alpha-methyl-5-HT maleate (alpha-m-5-HT), using the formalin test and the chronic constriction injury (CCI) model. An intrathecal catheter was implanted for injection of drugs. In the formalin test, flinches were counted from Minute 1 to 2 and Minute 5 to 6 (Phase 1) and then for 1-min periods at 5-min intervals from 10 to 60 min (Phase 2). In rats with CCI, hind paw withdrawal latency after thermal stimulation was measured. In the formalin test, intrathecal administration of alpha-m-5-HT (1 to 100 microg) dose-dependently suppressed the number of flinches in both Phases 1 and 2. In the CCI model, intrathecally administered alpha-m-5-HT (10 to 100 microg) attenuated thermal hyperalgesia in a dose-dependent manner. These effects were reversed by intrathecal pretreatment with a 5-HT(2A/2C) antagonist, ketanserin (30 microg), or a muscarinic receptor antagonist, atropine (30 microg). These findings suggest that spinal 5-HT(2A/2C) receptors mediate antinociception in inflammatory pain and neuropathic pain, and the muscarinic receptors contribute to this action. IMPLICATIONS: Activation of spinal 5-hydroxytryptamine(2A/2C) receptors mediate antinociception in rat-sustained pain models such as inflammatory pain and neuropathic pain, and spinal muscarinic receptors are involved in this action.     

A nonneuronal 5-hydroxytryptamine receptor 3 induces chloride secretion in the rat distal colonic mucosa

BACKGROUND: The 5-HT3 receptor is a serotonin receptor believed to reside on enteric neurons. However, several studies belie an exclusive neural localization. Our hypothesis is that the 5-HT3 receptor agonist, 2-methyl-5-HT (2Me5HT), induces chloride secretion despite neural blockade, which can be blocked by a 5-HT3 receptor antagonist. METHODS: Rat distal colon was stripped of its muscularis, mounted as mucosal sheets in Ussing chambers, and short-circuited. Adjacent sheets were pretreated with 1 micromol/L of the neurotoxin, tetrodotoxin, and incubated with 2Me5HT (50 micromol/L) alone or with a 5-HT3 (0.3 micromol/L ondansetron or 0.3 micromol/L tropisetron) or a 5-HT4 (0.3 micromol/L GR11808) receptor antagonist. Short-circuit current (I(sc)) was measured continuously. RESULTS: 2Me5HT caused an increase in I(sc), which was significantly (P <.01, repeated measures analysis of variance) inhibited by ondansetron (n = 8) and tropisetron (n = 5) but not by GR11808. CONCLUSIONS: A 5-HT3 receptor is present at the mucosal level that mediates chloride secretion by a nonneural pathway.     

Effect of different 5-hydroxytryptamine receptor subtype antagonists on the micturition reflex in rats

OBJECTIVE: To evaluate the effects of antagonists of different subfamilies of 5-hydroxytryptamine (5-HT) receptors on bladder function in anaesthetized and conscious rats. MATEERIALS AND METHODS: The urinary bladder of female anaesthetized rats was catheterized urethrally and filled with physiological saline until spontaneous bladder contractions occurred. Infravesical pressure was measured by a pressure transducer and displayed continuously on a chart recorder. The time of bladder quiescence (to the disappearance of rhythmic contractions) after injection with different compounds tested was recorded. Conscious rats underwent cystometry with chronically (infravesical) implanted catheters to continuously record bladder capacity (evaluated as amount of saline infused between voiding cycles) and maximal voiding pressure. The affinity for the human recombinant serotoninergic 5-HT1A subtype (inhibition of specific binding of [3H]8-hydroxy-2-(di-n-propylamino) tetralin) and the effects on the [35S]guanosine 5'-(gamma-thio) triphosphate (GTPgammaS) binding in HeLa cells was also evaluated. RESULTS: Among the compounds tested, only 4-(2'-methoxy-phenyl)-1-[2'-(n-2"-pyridinyl)-p-iodobenzamido]-ethyl-piperazine (p-MPPI) and methiothepin showed nanomolar affinity for the 5-HT1A receptors, the former being a neutral antagonist and the latter an inverse agonist in the [35S]GTPgammaS binding model. Intravenous injection of low doses of p-MPPI and methiothepin induced a dose-dependent disappearance of isovolumic bladder contractions in anaesthetized rats (> 10 min). At the highest doses, the dose-response curves were bell-shaped. The amplitude of bladder contractions was not markedly altered. The tested antagonists of 5-HT2, 5-HT3, 5-HT4, and 5-HT6 serotoninergic subtypes were poorly active or inactive in the model. Similarly, these compounds were inactive on cystometry in conscious rats, whereas p-MPPI and methiothepin induced a consistent increase in bladder capacity. Methiothepin also decreased the voiding pressure, whereas p-MPPI did not affect this variable. CONCLUSIONS: These findings confirm that only selective 5-HT1A receptor antagonists have favourable effects on the bladder, inducing an increase in bladder capacity with no derangement of bladder contractility.     

Influence of 5-hydroxytryptamine and the effect of a new serotonin receptor antagonist (sarpogrelate) on detrusor smooth muscle of streptozotocin-induced diabetes mellitus in the rat.

BACKGROUND: We evaluated the changes in the response of detrusor muscle to 5-hydroxytryptamine (5-HT; serotonin) and its receptor mechanisms in pathologic bladder of diabetes mellitus rats. METHODS: Eight-week-old male Wistar rats were given an intraperitoneal injection of streptozotocin (STZ, 60 mg/kg) in order to induce diabetes mellitus (4 weeks' duration). The bladder strips (10 x 2 mm) were subjected to a tension of 1.0 g in organ baths for the measurements of isotonic contractile responses to 5-HT or relaxation responses to 5-HT antagonists. RESULTS: Bladder strips from STZ-diabetic rats responded to 5-HT with greater contractions than did those of control rats, given vehicle alone. However, bladder strip weights gradually increased in the STZ-diabetic rats as compared with control rats and when the contractile response to 5-HT was expressed as a percentage of the response to 10(-1) mol/L KCl there was no statistically significant difference between the groups. In both control and diabetic rats, the 5-HT3 selective antagonist did not markedly inhibit the contractile response to 5-HT. In contrast, the 5-HT2A selective antagonist inhibited the contractile response to 5-HT in a dose-dependent manner. CONCLUSIONS: These results suggest that the increased contractile response to 5-HT in diabetic rats' bladder is related to smooth muscle hypertrophy and/or hyperplasia and indicate that this effect is mediated by activation of 5-HT2A receptors.     

Protective effect of a new C5a receptor antagonist against ischemia-reperfusion injury in the rat small intestine

BACKGROUND: The complement system is a major contributor to the pathogenesis of intestinal ischemia-reperfusion (I/R) injury. We have studied the action of an orally active complement factor 5a (C5a) receptor antagonist, the cyclic peptide AcF-(OPdChaWR) [Ac-Phe(Orn-Pro-d-cyclohexylalanine-Trp-Arg)] against local and remote intestinal I/R injuries in rats. MATERIALS AND METHODS: Anesthetized rats were administered with AcF-(OPdChaWR) at doses of 1 mg/kg intravenously or 0.3, 1, or 10 mg/kg orally with pyrogen-free saline for sham control animals. The superior mesenteric artery was occluded for 30 min and the intestine reperfused for 120 min. Changes associated with tissue injury were assessed by neutropenia, intestinal edema, serum tumor necrosis factor-alpha, serum haptoglobin, plasma aspartate aminotransferase, and histopathology. RESULTS: Pretreatment with either a single intravenous dose (1 mg/kg), or a single oral dose (10 mg/kg) of AcF-(OPdChaWR) significantly inhibited I/R induced neutropenia, the elevated serum levels of tumor necrosis factor-alpha, haptoglobin, and plasma aspartate aminotransferase, as well as intestinal edema. Histological analysis of AcF-(OPdChaWR)-treated I/R animals showed markedly reduced mucosal layer damage compared to that of untreated rats. CONCLUSIONS: These results indicate that a potent antagonist of C5a receptors on human cells protects the rat small intestine from I/R injury after oral or intravenous administration. Small molecule C5a antagonists may have some therapeutic utility in reperfusion injury.     

Portal serotonin infusion and glucose disposal in conscious dogs

Whether serotonin (5-hydroxytryptamine [5-HT]) enhances net hepatic glucose uptake (NHGU) during glucose infusion was examined in conscious 42-h-fasted dogs, using arteriovenous difference and tracer ([3-(3)H]glucose) techniques. Experiments consisted of equilibration (-120 to -30 min), basal (-30 to 0 min), and experimental (0-390 min) periods. During the experimental period, somatostatin, fourfold basal intraportal insulin, basal intraportal glucagon, and peripheral glucose (to double the hepatic glucose load) were infused. In one group of dogs (SER; n = 8), saline was infused intraportally from 0 to 90 min (P1), and 5-HT was infused intraportally at 10, 20, and 40 microg.kg(-1).min(-1) from 90 to 150 (P2), 150 to 210 (P3), and 210 to 270 (P4) min, respectively. In the other group (SAL; n = 8), saline was infused intraportally from 0 to 270 min. NHGU in SAL was 12.4 +/- 2.3, 14.9 +/- 2.7, 13.4 +/- 2.1, and 15.1 +/- 1.8 micromol.kg(-1).min(-1) in P1 to P4, respectively, whereas NHGU in SER averaged 13.2 +/- 3.0, 16.4 +/- 2.4, 19.0 +/- 2.4 (P < 0.05 vs. SAL), and 22.0 +/- 2.9 micromol.kg(-1).min(-1) (P < 0.05 vs. SAL). Nonhepatic glucose uptake ( micromol.kg(-1).min(-1)) in SAL was 31.7 +/- 4.9, 43.9 +/- 5.1, 55.1 +/- 5.6, and 66.2 +/- 8.6 during P1 to P4, respectively, whereas in SER, the corresponding values were 26.1 +/- 5.7, 31.6 +/- 9.4, 35.1 +/- 7.6 (P < 0.05 vs. SAL), and 34.7 +/- 7.7 (P < 0.05 vs. SAL). Intraportal 5-HT enhances NHGU but blunts nonhepatic glucose uptake, raising the possibility that hepatic-targeted 5-HT or 5-HT receptor agonists might reduce postprandial hyperglycemia.     

D1-like dopamine receptor activation and natriuresis by nitrocatechol COMT inhibitors

BACKGROUND: In recent years, several nitrocatechol derivatives (tolcapone, entacapone, and nitecapone) have been developed and found to be highly selective and potent inhibitors of catechol-O-methyltransferase (COMT). More recently, natriuretic properties were described for two of these compounds (entacapone and nitecapone), although this was not accompanied by enhanced urinary excretion of dopamine. We hypothesized that nitrocatechol derivatives stimulate D1-like dopamine receptors. METHODS: Adult male Wistar rats were treated with a nitrocatechol COMT inhibitor (entacapone, tolcapone, or nitecapone, 30 mg/kg, orally), and the urinary excretion of dopamine and sodium was quantitated. The interaction of nitrocatechol derivatives with D1-like receptors was evaluated by their ability to displace [3H]-Sch23390 binding from membranes of rat renal cortex and cAMP production in opossum kidney (OK) cells. RESULTS: Urinary excretion of sodium (micromol/h) was markedly increased by all three nitrocatechol derivatives: vehicle, 55.0 +/- 5.6; entacapone, 98.4 +/- 9.3; tolcapone, 97.5 +/- 9.3; and nitecapone, 120.5 +/- 12.6. Pretreatment with the selective D1 antagonist Sch 23390 (60 microg/kg) completely prevented their natriuretic effects. Nitecapone and tolcapone were equipotent (IC50s of 48 and 42 micromol/L) and more potent than entacapone and dopamine (IC50s of 107 and 279 micromol/L) in displacing [3H]-Sch23390 binding. In OK cells, all three nitrocatechol derivatives significantly increased cAMP accumulation and reduced Na(+)/H(+) exchange and Na(+),K(+)-ATPase activities, this being prevented by a blockade of D1-like receptors. CONCLUSION: Stimulation of D1-like dopamine receptors and inhibition of Na(+)/H(+) exchange and Na(+),K(+)-ATPase activities by nitrocatechol COMT inhibitors may contribute to natriuresis produced by these compounds.     

Characterization of intracavernous pressure increase induced by Ym348, a novel 5-HT2C receptor agonist, in anesthetized rats

PURPOSE: We investigated the effects of the novel 5-HT2C receptor agonist YM348 [(S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine] on ICP in anesthetized rats and clarified whether behavioral changes such as hypolocomotion induced at a high dose are a cause of the inverted U-shaped PE dose-response curves in conscious rats. MATERIALS AND METHODS: Male Wistar rats (Japan SLC, Shizuoka, Japan) weighing 250 to 315 gm were used. The pro-erectile effect of YM348 (2.03 to 67.7 microg/kg subcutaneously) was examined in conscious rats. ICP was also monitored after YM348 treatment (0.677 to 67.7 mug/kg subcutaneously) in anesthetized rats. Response number, latency, duration, peak pressure and area under the curve were measured. The selective 5-HT2C receptor antagonist SB242084 [(6-chloro-5-methyl-1-[6-(2-methylpyridin-3-yloxy) pyridin-3-yl carbamoyl] indoline)] (0.03 to 1 mg/kg intraperitoneally) was administered 30 minutes before YM348 treatment. RESULTS: YM348 induced PE and ICP increases, and was significantly inhibited by SB242084. Dose-response curves for the number of PEs and ICP increases were an inverted U shape. YM348 decreased the latency of but did not affect the quality of ICP (duration, peak pressure and area under the curve) even at the highest dose. CONCLUSIONS: Activation of 5-HT2C receptor increased ICP and, as a result, induced PE. Since the dose-response curve for the number of ICP increases under anesthetized, behavior independent conditions still showed an inverted U shape, behavioral changes were not likely to have contributed to the inverted U-shaped dose-response curves for the number of PEs in conscious rats. Furthermore, the certain magnitude of ICP increases was likely to have occurred despite the stimulus intensity after the level of 5-HT2C receptor activation crossed the threshold.     

Therapeutic benefits of 5-hydroxytryptamine inhibition following pulmonary embolism.

The smooth muscle-constricting, platelet amine, serotonin (5-hydroxytryptamine; 5-HT) is theorized to play an important role in the cardiopulmonary dysfunction that accompanies embolization. The present study was designed to examine this hypothesis. Autologous clot, 0.75 g/kg, was injected IV into 14 dogs. After 30 minutes, one half of the animals were randomly assigned to the treatment group and received a bolus infusion of 0.15 mg/kg ketanserin, a quinazoline derivative known to be a selective 5-HT receptor antagonist. Five minutes after embolization there were increases in mean pulmonary arterial pressure (MPAP) from 12 mm to 48 mmHg (p less than 0.001); pulmonary vascular resistance (PVR) from 2.2 mm to 12.2 mmHg X min/L (p less than 0.001); physiologic shunt (QS/QT) from 12% to 44% (p less than 0.01); and physiologic dead space (VD/VT), calculated from end tidal and arterial PCO2, from 8% to 39% (p less than 0.001). Within 15 minutes platelet counts decreased from 186,000/mm3 to 134,800/mm3 (p less than 0.05); 5-HT contained in circulating platelets fell from 1.71 micrograms/ to 1.44 micrograms/10(9) platelets (p less than 0.05). Five minutes after ketanserin, MPAP declined to 27 mmHg and was lower than the control value of 41 mmHg (p less than 0.05); PVR decreased to 6.2 mmHg X min/L, lower than 12 mmHg X min/L in controls (p less than 0.01); QS/QT fell to 26% in contrast to 47% in controls (p less than 0.05); and VD/VT declined moderately to 32% (p less than 0.05), although this value was not different from 38% in control animals. Cardiopulmonary function continued to improve in treated animals until termination of the experiment at four hours when pulmonary angiograms and perfusion scans demonstrated vascular recruitment compared with untreated embolized control dogs. These data demonstrate that the cardiopulmonary consequences of experimental embolization are primarily determined by the vasoconstrictive and bronchoconstrictive actions of 5-HT.     

Remifentanil preconditioning confers delayed cardioprotection in the rat

BACKGROUND: Preconditioning with remifentanil (RPC) provides immediate cardioprotection in rats via all three types of opioid (OP) receptor. This study sought to investigate whether remifentanil also confers delayed cardioprotection via OP receptors. METHODS: Male rats received preconditioning either by ischaemia (IPC; 5 min occlusion, 5 min reperfusion x 3) or with remifentanil (RPC; 1, 5, 10, and 20 microg kg(-1) min(-1), 20 min infusion). After 24 h, all animals were subjected to 30 min occlusion of the left coronary artery and 2 h of reperfusion. Subsequently, the time-course effect of RPC (10 microg kg(-1) min(-1), 20 min infusion) was determined at 12, 16, 24, 32, 36, and 48 h intervals, using the same experimental procedure. The effect of RPC (10 microg kg(-1) min(-1), 20 min infusion) and IPC in the presence of selective OP receptor antagonists was evaluated at the 24 h interval. Infarct size (IS), as a percentage of the area at risk (AAR), was determined. RESULTS: Pre-treatment with remifentanil at 1, 5, 10, and 20 microg kg(-1) min(-1) significantly reduced the IS/AAR at 24 h with the maximum effect at 10 microg kg(-1) min(-1). Remifentanil at 10 microg kg(-1) min(-1) significantly reduced the IS at 12 h [32.5 (sd 9.1)%]; 16 h [26.1 (2.8)%]; 24 h [19.5 (5.0)%]; 32 h [31.2 (9.1)%]; and 36 h [36.4 (9.4)%] after drug administration. The maximal reduction in IS was seen at 24 h and the effect completely disappeared at 48 h [36.4 (9.4)%]. The protective effect of RPC was abolished or significantly attenuated by blockade of any of the three OP receptors with selective antagonists. CONCLUSIONS: Like IPC, remifentanil produces delayed cardioprotection in anaesthetized rats 12-36 h after administration. The protective effect is mediated via all three OP receptors.     

Protective effects of a potent C5a receptor antagonist on experimental acute limb ischemia-reperfusion in rats

OBJECTIVES: The capacity of a potent C5a receptor antagonist to inhibit various parameters of local and remote organ injury following lower limb ischemia-reperfusion (I/R) in rats was investigated. METHODS: Rats were subjected to 2 h bilateral hindlimb ischemia and 4 h reperfusion. Drug-treated rats received AcF-[OPdChaWR] (1 mg/kg) iv either 10 min before ischemia or 10 min prior to reperfusion, or orally (10 mg/kg) 30 min prior to ischemia. Levels of circulating creatine kinase (CK), lactate dehydrogenase (LDH), alanine and aspartate aminotransferase (ALT/AST), creatinine, blood urea nitrogen (BUN), polymorphonuclear leukocytes (PMNs), and calcium (Ca(++)) and potassium (K(+)) ions were determined. Other parameters measured included urinary protein levels, muscle edema, and myeloperoxidase (MPO) concentrations in the lung, liver, and muscle along with liver homogenate tumor necrosis factor-alpha (TNF-alpha) concentrations.L RESULTS: imb I/R injury was characterized by significant elevations of CK, LDH, ALT, AST, creatinine, BUN, proteinuria, PMNs, serum K(+), muscle edema, organ MPO, and liver homogenate TNF-alpha concentrations, but a significant reduction in serum Ca(2+) concentrations. When rats were treated with AcF-[OPdChaWR], there were significant improvements in all these parameters. CONCLUSIONS: These results indicate a pivotal role for C5a in inducing local and remote organ injury and suggest a possible new drug therapeutic category for preventing anticipated tissue injury associated with I/R.     

Independent regulation of in vivo insulin action on glucose versus K(+) uptake by dietary fat and K(+) content

Insulin stimulates both glucose and K(+) uptake, and high-fat feeding is known to decrease insulin-stimulated glucose uptake. The purpose of this study was to examine whether insulin's actions on glucose and K(+) uptake are similarly decreased by a high-fat diet. Wistar rats were fed a standard control (12.2% fat; n = 6) or high-fat (66.5% fat; n = 13) diet for 15 days. Because K(+) content was 1% in the control and 0.5% in the high-fat diet and because the rats ate less of the high-fat diet, we also compared the high-fat diet with 0.5% K(+) (HFD; n = 7) to a high-fat diet supplemented with 1.5% K(+) (HFD+K; n = 6). K(+) intake was matched between the control and HFD+K groups (246 +/- 8 vs. 224 +/- 2 mg/day), but was lower in the HFD group (78 +/- 10 mg/day; P < 0.05). Insulin-stimulated glucose and K(+) uptake were determined by hyperinsulinemic (5 mU.kg(-1).min(-1)) glucose and K(+) clamps. The HFD depressed both insulin-stimulated glucose uptake compared to the control (133 +/- 5 vs. 166 +/- 7 micromol.kg(-1).min(-1); P < 0.05) and K(+) uptake (5.5 +/- 0.9 vs. 8.9 +/- 1.0 micromol.kg(-1).min(-1); P < 0.05) compared to the control. However, insulin-stimulated K(+) uptake was unchanged in the HFD+K versus in the control group (10.0 +/- 0.6 vs. 8.9 +/- 1.0 micromol.kg(-1).min(-1); P > 0.05), whereas insulin-stimulated glucose uptake in the HFD+K group was decreased to a rate (137 +/- 9 micromol.kg(-1).min(-1)), similar to that of the HFD group. We concluded that the decrease in insulin-stimulated K(+) uptake during high-fat feeding was a result of decreased K(+) intake, and that insulin's actions on glucose uptake and K(+) uptake are independently regulated by dietary fat and K(+) content, respectively.     

Production of cysteinyl-dopamine during intravenous dopamine therapy

BACKGROUND: Oxidized dopamine rapidly forms thiol-conjugates with --SH groups on cysteine, glutathione, and proteins. We used cysteinyl-dopamine production as an index of thioester production during intravenous dopamine treatment of critically ill patients. METHODS: Cysteinyl-dopamine and catecholamines were measured by high-performance liquid chromatography with electrochemical detection. The production of cysteinyl-dopamine by purified human neutrophils was measured using dopamine (1 micromol/L) and cysteine (1 mmol/L) concentrations similar to those found during dopamine treatment. To examine the impact of endotoxic shock on cysteinyl-dopamine production, anesthetized rats were given dopamine (12 to 15 microg/kg/min intravenously) with or without endotoxin (50 mg/kg intravenously). RESULTS: In vitro, neutrophils converted 26% of dopamine to cysteinyl-dopamine (30 min at 37 degrees C). Activating neutrophils with zymogen increased dopamine consumption from 26 to 68%, but only 36% appeared as cysteinyl-dopamine. The remainder may have been oxidized to other cysteinyl derivatives. Endotoxin increased cysteinyl-dopamine in rat plasma from 2.5 nmol/L (range <0.2 to 11) to 9.7 nmol/L (range <0.3 to 31, P = 0.1). After four hours, with or without endotoxin, cysteinyl-dopamine was <0.3 nmol/L in cerebrospinal fluid. In the plasma of eight patients receiving dopamine (6 to 20 microg/kg/min for 1 to 3 days), dopamine was 0.5 to 9.9 micromol/L, and cysteinyl-dopamine was 48 to 1660 nmol/L. Cysteinyl-dopamine was 4.3 to 22.6% of dopamine and correlated with leukocyte count (r(2) = 0.388, P = 0.099). CONCLUSIONS: A significant fraction of exogenously administered dopamine reacts with -SH groups of cysteine and probably also with -SH groups on peptides and proteins. During brief dopamine treatment of endotoxic shock in rats, neither dopamine nor cysteinyl-dopamine crossed the blood-brain barrier.     

A small molecule C5a receptor antagonist protects kidneys from ischemia/reperfusion injury in rats

BACKGROUND: C5a has been implicated in numerous pathophysiological conditions, including ischemia/reperfusion (I/R) injury of the kidney. We examined whether a novel and specific C5a receptor antagonist, the cyclic compound AcF-[OPdChaWR] could moderate I/R-induced renal injury in rats. METHODS: Female Wistar rats were subjected to renal ischemia (60 min) and reperfusion (5 h). Rats were treated with either 1 mg/kg IV in 5% ethanol/saline or 10 mg/kg PO in 25% ethanol/saline prior to ischemia. I/R injury was characterized by significant tissue hemorrhage with increased microvascular permeability, elevated renal tissue levels of tumor necrosis factor-alpha (TNF-alpha) and myeloperoxidase (MPO), increased serum levels of creatinine and aspartate aminotransferase (AST) and hematuria. RESULTS: Pre-ischemic treatment with the C5a receptor (C5aR) antagonist (1 mg/kg IV or 10 mg/kg PO) substantially inhibited or prevented I/R-induced hematuria, vascular leakage, tissue levels of TNF-alpha and MPO, and serum levels of AST and creatinine. Histological examination of kidneys from antagonist pretreated I/R animals showed a marked reduction in tissue damage compared to drug-free I/R rats. This antagonist, however, did not inhibit complement-mediated lysis of red blood cells, suggesting unimpaired formation of the membrane attack complex (MAC). CONCLUSIONS: The results demonstrate for the first time that a selective antagonist of both human and rat C5a receptors, given either intravenously or orally, significantly protects the kidney from I/R injury in the rat. We conclude that C5a is an important pathogenic agent in renal I/R injury, and that C5a receptor antagonists may be useful therapeutic agents for the pretreatment of anticipated renal reperfusion injury in humans.     

Cariporide (HOE 642) attenuates leukocyte activation in ischemia and reperfusion.

Cariporide (HOE 642) ameliorates myocardial ischemia/reperfusion (I/R) injury, by the well established reduction of cytosolic [Ca(2+)] in cardiac myocytes through inhibition of Na(+)/H(+) exchange. However, postischemic inflammation also contributes to I/R injury. We tested the hypothesis that cariporide also modulates the inflammatory response. The effect of cariporide on L-selectin expression by human leukocytes in vitro and leukocyte adhesion and emigration in the reperfused rat cremaster muscle in vivo were studied. The rat cremaster muscle was exteriorized for intravital videomicroscopy, induction of ischemia (90 min), and reperfusion (90 min). Eleven rats were pretreated with cariporide (9 mg/kg body weight IV) whereas 11 rats received saline. Leukocyte adhesion was quantified offline. Human venous blood was incubated with cariporide (3 micromol/L) or saline, stimulated with formyl- methionine-leucine-phenylalanine (10(-10)-10(-6) mol/L), and granulocyte L-selectin expression was analyzed by flow cytometry. Cariporide reduced leukocyte rolling and adhesion by approximately 35% and 45%, respectively, after 30 min of reperfusion. Leukocyte extravasation was decreased by approximately 85% after 90 min. Cariporide increased L-selectin shedding at each formyl-methionine-leucine-phenylalanine concentration, reducing the 50% effective dose from 9.95 to 4.68 nmol/L. Thus, cariporide may ameliorate I/R injury not only by the known reduction of cytosolic [Ca(2+)] in cardiomyocytes, but also by attenuating leukocyte-dependent inflammatory responses. Promotion of L-selectin shedding from activated leukocytes may present a mechanism underlying this newly detected effect. IMPLICATIONS: This study provides evidence that inhibition of Na(+)/H(+) exchange by cariporide (HOE 642) attenuates the postischemic inflammatory response. Leukocyte adhesion and emigration, assessed by in vivo microscopy, were markedly reduced in rat cremaster muscle, possibly because of increased L-selectin shedding of activated leukocytes as demonstrated by flow cytometry.     

吗啡预处理减轻小鼠海马神经元氧-糖缺失/恢复损伤的影响因素

目的 探讨吗啡预处理减轻小鼠海马神经元氧-糖缺失/恢复损伤的影响因素(剂量和作用时间窗).方法 急性分离小鼠海马组织,制备脑片,行4部分实验,实验Ⅰ:应用吗啡0.1、0.3、0.5、1.0、3.0、10.0 μmol/L孵育脑片30 min,再常规培养30 min,缺氧无糖20 min,复氧复糖2 h.实验Ⅱ:应用吗啡3.0 μmol/L孵育脑片5、15、30、45、60 min,再常规培养30 min,缺氧无糖20 min,复氧复糖2 h.实验Ⅲ:应用吗啡3.0 μmol/L孵育脑片30 min,再常规培养即刻、5、15、30、60、120 min时行缺氧无糖20 min,复氧复糖2 h.实验Ⅳ:应用蛋白激酶C(PKC)非特异性抑制剂白屈菜赤碱10 μmol/L、选择性新奇型PKCε(nPKCε)抑制剂εVI.2 2 μmol/L、特异性钙-钙调蛋白依赖性蛋白激酶Ⅱ(CaMKⅡ)抑制剂AIP 2 μmol/L、N.甲基-D-天冬氨酸(NMDA)受体特异性阻断剂MK-801 10 μmol/L孵育脑片30 min,再与吗啡3.0 μmol,L共同孵育30 min,常规培养30 min,缺氧无糖20 min,复氧复糖2 h.计算神经元存活率.结果 与氧-糖缺失/恢复组比较,吗啡0.5、1.0、3.0、10.0 μmol/L预处理组、吗啡预处理15、30、45、60 min组、吗啡预处理后常规培养即刻、5、15、30、60 min组神经元存活率升高(P<0.05).PKC、nPKGt、CaMKⅡ抑制剂和NMDA受体阻断剂可部分抑制吗啡预处理升高神经元存活率的作用.结论 有效减轻小鼠海马神经元氧-糖缺失,恢复损伤的吗啡预处理方式为:浓度0.5～1.0 μmol/L,持续时间15～60 min,间隔时间小于60 min;其保护作用可能部分依赖于PKC、nPKCε、CaMKⅡ的活化和NMDA受体的激活.     

5-HT5A受体在长春新碱致神经病理性痛大鼠脊髓背角星形胶质细胞活化中的作用

目的 评价5-羟色胺5A受体(5-HT5AR)在长春新碱致神经病理性痛大鼠脊髓背角星形胶质细胞活化中的作用.方法 雄性成年SD大鼠40只,体重180～200 g,随机分为4组(n=10):对照组(C组)、神经病理性痛组(P组)、空载体腺病毒组(B组)和siRNA重组腺病毒载体组(S组).C组腹腔注射生理盐水1 ml;P组、B组和S组第1～5天和第8～12天每天定时腹腔注射0.1 mg/kg长春新碱建立大鼠神经病理性痛模型.腹腔给药结束第2天测定机械痛阈,然后P组、B组和S组分别鞘内注射人工脑脊液、空载体腺病毒和siRNA重组腺病毒载体25μl.鞘内给药后第7天测定机械痛阈,然后处死大鼠,取L4.5脊髓组织,测定脊髓背角5-HT5AR及胶原纤维酸性蛋白(GFAP)的表达.结果 与C组比较,P组、B组和S组各时点机械痛阈降低,脊髓背角5-HT5AR和GFAP的表达均上调(P＜0.05);与P组比较,S组鞘内给药后第7天机械痛阈降低,脊髓背角5-HT5AR表达下调,GFAP表达上调(P＜0.05),B组上述指标差异无统计学意义(P＞0.05).结论 5-HT5AR参与了星形胶质细胞活化的抑制过程,从而减轻长春新碱致大鼠神经病理性痛.     

The effects of ketamine and its enantiomers on the morphine- or dexmedetomidine-induced antinociception after intrathecal administration in rats

BACKGROUND: The spinal administration of some N-methyl-d-aspartate receptor antagonists results in antinociception and potentiates the effects of opioids and alpha2-adrenoceptor agonists, but ketamine and its enantiomers have not been examined. The present study investigated the interactions of racemic ketamine, R(-)-ketamine and S(+)-ketamine with morphine and with dexmedetomidine. METHODS: Intrathecal catheters were implanted into male Wistar rats. Three days later, the acute nociceptive sensitivity was assessed using the tail-flick test. Analgesic latencies were converted to the percentage maximum possible effect. The dose that yielded 50% of the maximum possible effect (ED50) and dose-response and time-course curves were determined for the ketamines (30-300 microg), morphine (0.1-3.0 microg), dexmedetomidine (0.3-10.0 microg), and mixtures of two doses of ketamines (30 or 100 microg) with different doses of morphine or dexmedetomidine for fixed-dose analysis. RESULTS: Neither racemic ketamine nor its enantiomers alone had a significant effect on the tail-flick test, with the exception of the highest dose of racemic ketamine, which caused motor impairment. Morphine and dexmedetomidine each produced dose-dependent antinociception, with ED50 of 1.7 microg (95% confidence interval: 1.04-2.32) and 4. 85 microg (3.96-5.79), respectively. A low dose (30 microg) of racemic ketamine or its enantiomers did not influence the ED50 of morphine significantly. Coadministration of 100 microg racemic ketamine or S(+)-ketamine, but not R(-)-ketamine, significantly enhanced and prolonged the antinociceptive effect of morphine. Both doses of racemic ketamine or its isomers significantly decreased the ED50 value for dexmedetomidine, although the higher dose of racemic or S(+)-ketamine had the highest potency. One-hundred micrograms of racemic ketamine or S(+)-ketamine also prolonged the effects of dexmedetomidine. CONCLUSIONS: These data indicate that racemic ketamine and S(+)-ketamine, but not R(-)-ketamine, exhibit similar effectiveness in potentiating the antinociceptive effects of both morphine and dexmedetomidine.     

Effect of three different doses of urapidil on blood glucose concentrations in the streptozotocin diabetic rat

BACKGROUND AND OBJECTIVE: Diabetes mellitus associated with hypertension often causes perioperative complications. The alpha1-adrenoceptor antagonist/5-hydroxytryptamine-1A receptor agonist urapidil is an approved drug used in hypertension and hypertensive emergencies. 5-Hydroxytryptamine-1A (5-HT1A) receptor agonists impair glucose metabolism. To evaluate a possible dose-dependent hyperglycaemic effect of urapidil due to its 5-HT1A receptor agonistic properties, the effect of three doses of urapidil on hyperglycaemia in the streptozotocin diabetic rat was investigated. METHODS: Male Wistar-Kyoto rats were made diabetic by streptozotocin and randomly allocated to the following daily treatments for 7 days (n = 6 each): urapidil 6 mg kg(-1), urapidil 20 mg kg(-1), urapidil 60 mg kg(-1), insulin 4 IU kg(-1) subcutaneously. One diabetic group and one non-diabetic healthy group served as controls. RESULTS: Treatment for 7 days with urapidil 20 mg kg(-1) and urapidil 60 mg kg(-1) reduced mean glucose concentrations significantly (urapidil-20: 15.6 +/- 1.1 mmol L(-1), P = 0.023; urapidil-60: 15.8 +/- 0.8 mmol L(-1), P = 0.04) compared with diabetic controls (20.9 +/- 0.8 mmol L(-1)), whereas those after urapidil 6 mg kg(-1) were similar to diabetic controls. Insulin treatment normalized blood glucose concentrations. CONCLUSIONS: The alpha1-adrenoceptor antagonist/5-HT1A receptor agonist urapidil has no hyperglycaemic effect on experimental diabetes mellitus, even in high doses, despite its 5-HT1A receptor agonistic properties.     

High-dose S(+)- ketamine improves neurological outcome following incomplete cerebral ischemia in rats

PURPOSE: To determine the effects of the non-competitive NMDA-receptor antagonist S(+)-ketamine on neurological outcome in a rat model of incomplete cerebral ischemia. METHODS: Thirty rats were anesthetized, intubated and mechanically ventilated with isoflurane, O2 30% and nitrous oxide 70%. Following surgery animals were randomly assigned to one of the following treatment groups: Rats in group 1 (n = 10,OFF control) received fentanyl (bolus: 10 microg x kg(-1) i.v.; infusion 25 microg x kg(-1) x h(-1)) and N2O 70% / O2. Rats in group 2 (n = 10) received O2 30% in air and low-dose S(+)-ketamine (infusion: 0.25 mg x kg(-1) x min(-1)). Rats in group 3 (n = 10) received O2 30% in air and high-dose S(+)-ketamine (infusion: 1.0 mg x kg(-1) min(-1)). Following 30 min equilibration period ischemia was induced by combined unilateral common carotid artery ligation and hemorrhagic hypotension to 35 mm Hg for 30 min. Plasma catecholamines were assayed before and at the end of ischemia. Neurological deficit was evaluated for three postischemic days. RESULTS: Neurological outcome was improved with high-dose S(+)-ketamine when compared to fentanyl / N2O -anesthetized controls (9 vs. 1 stroke related deaths, P<0.05). Increases in plasma catecholamine concentrations were higher in fentanyl / N2O -anesthetized (adrenaline baseline 105.5+/-92.1 pg x ml(-1), during ischemia 948+/-602.8 pg x ml(-1), P<0.05; noradrenaline baseline 407+/-120.2 pg x ml(-1), ischemia 1267+/-422.2 pg x ml(-1), P <0.05) than in high-dose S(+)-ketamine-treated animals (adrenaline baseline 71+/-79.5 pg x ml(-1), ischemia 237 +/-131.9; noradrenaline baseline 317.9+/-310.5 pg x ml(-1), ischemia 310.5+/-85.7 pg x ml(-1)). CONCLUSION: Neurological outcome is improved following incomplete cerebral ischemia with S(+)-ketamine. Decreases in neuronal injury may be related to suppression of sympathetic discharge.