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Intrathecal (i.t.) administration into mice of N-ethylmaleimide (NEM), a cysteine protease inhibitor, produced a characteristic behavioral response, the biting and/or licking of the hindpaw and the tail along with slight hindlimb scratching directed toward the flank. The behavior induced by NEM was inhibited by the intraperitoneal injection of morphine. We have recently reported that dynorphin A and, more potently big dynorphin, consisting of dynorphins A and B, produce the same type of nociceptive response whereas dynorphin B does not [Tan-No K, Esashi A, Nakagawasai O, Niijima F, Tadano T, Sakurada C, Sakurada T, Bakalkin G, Terenius L, Kisara K. Intrathecally administered big dynorphin, a prodynorphin-derived peptide, produces nociceptive behavior through an N-methyl-d-aspartate receptor mechanism. Brain Res 2002;952:7-14]. The NEM-induced nociceptive behavior was inhibited by pretreatment with dynorphin A- or dynorphin B-antiserum and each antiserum also reduced the nociceptive effects of i.t.-injected synthetic big dynorphin. The characteristic NEM-evoked response was not observed in prodynorphin knockout mice. Naloxone, an opioid receptor antagonist, had no effects on the NEM-induced behavior. Ifenprodil, arcaine and agmatine, antagonists at the polyamine recognition site on the N-methyl-D-aspartate (NMDA) receptor ion-channel complex, and MK-801, an NMDA ion-channel blocker inhibited the NEM-induced effects. Ro25-6981, an antagonist of the NMDA receptor subtype containing NR2B subunit was not active. NEM completely inhibited degradation of dynorphin A by soluble and particulate fractions of mouse spinal cord. Collectively, the results demonstrate that endogenous prodynorphin-derived peptides are pronociceptive in uninjured animals, and required for the NEM-induced behavior. The NEM effects may be mediated through inhibition of the degradation of endogenous dynorphins, presumably big dynorphin that in turn activates the NMDA receptor ion-channel complex by acting on the polyamine recognition site.

To investigate the effect of angiotensin (ANG) II on aldosterone (ALDO) secretion, we measured arterial and adrenal venous plasma aldosterone concentrations in anesthetized dogs. The intraadrenal arterial infusion of ANG II (0.3 ng/kg/min) or potassium chloride (KCl) (0.6 mg/min) increased ALDO secretion. The changes in ALDO secretion in response to ANG II were tested during the concomitant arterial infusion of two graded doses of losartan (10 and 100 ng/kg/min), PD 123319 (50 and 500 ng/kg/min), nifedipine (25 and 250 ng/kg/min), or TMB-8 (2 and 20 microg/kg/min). All of these test drugs except PD123319 inhibited the ANG II-induced increase in ALDO secretion. Losartan did not affect the KCl-induced increase in ALDO secretion. These results indicate that ANG II acts on ANG II type 1 receptors in the adrenal gland and enhances ALDO secretion. They also suggest the involvement of both intracellular and extracellular calcium in the aldosterone response to stimulation by ANG II. Under these in vivo experimental conditions, the KCl-stimulated ALDO secretion does not appear to involve ANG II formation in the adrenal gland.

Although the renal nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) system plays an important role in maintaining urinary sodium and water excretion, effects of an authentic NO donor sodium nitroprusside (SNP) on urine formation have been controversial. In this study, we examined whether SNP increases renal NO release and cGMP production and induces natriuresis in the denervated kidney of anesthetized dogs. The intrarenal arterial infusion of SNP at 10, 30, and 100 ng/kg/min did not affect renal function or NO-cGMP production. The higher dose of SNP (1.000 ng/kg/min) reduced systemic blood pressure and urine flow rate. The antidiuresis was observed also in the contralateral control kidney, the degree of which was larger than that observed in the ipsilateral SNP-infused kidney. During the SNP infusion, reductions in urinary Na + excretion, fractional Na + excretion. and urinary nitrite + nitrate excretion occurred in the control kidney but not in the SNP-infused kidney. Urinary cGMP excretion and renal venous plasma cGMP concentration were significantly increased during the SNP infusion in the SNP-infused kidney but not in the control kidney. These renal effects of SNP were similar to those obtained by intrarenal arterial infusion of a specific NO donor, NOC 7 (300 ng/kg/min). These results suggest that SNP can produce nitric oxide and increase cGMP levels in the kidney and suppress sodium reabsorption. but the natriuretic property of SNP may be masked by its counteracting effects including the systemic hypotension in anesthetized dogs.

We investigated the effect of cilnidipine, a novel dihydropyridine Ca2+ channel blocker possessing blocking actions on N-type and L-type voltage-dependent Ca2+ channels (VDCCs), in comparison with the L-type VDCC blocker nifedipine, on adrenal catecholamine secretion in response to splanchnic nerve stimulation (SNS), acetylcholine (ACh), the nicotinic receptor stimulant 1,1-dimethyl-4-phenyl-piperazinium (DMPP), and muscarine in anesthetized dogs. Ca2+ channel blockers and cholinergic agonists were infused and injected, respectively, into the adrenal gland through the phrenicoabdominal artery. Cilnidipine (0.3-3 microg/min) inhibited increases in both epinephrine (EPI) and norepinephrine (NE) output induced by SNS (2 Hz), ACh (1.5 microg), and DMPP (0.2 microg). However, cilnidipine inhibited increase in NE output induced by muscarine (1 microg) without affecting increase in EPI output. Nifedipine (0.3-3 microg/min) inhibited the ACh- and DMPP-induced increases in EPI and NE output without affecting the SNS- and muscarine-induced increases in EPI and NE output. From these results, it seems likely that the inhibition by cilnidipine of the SNS-induced EPI and NE secretion and of the muscarine-induced NE secretion is related to its blocking action on N-type VDCCs.

We examined the effects of glucagon on the release of adrenal catecholamines (CAs) in response to splanchnic nerve stimulation (SNS), acetylcholine (ACh), the nicotinic receptor stimulant 1,1-dimethyl-4-phenyl-piperazinium iodide (DMPP), and muscarine in anesthetized dogs. Glucagon and the cholinergic drugs were infused and injected, respectively, into the adrenal gland through the phrenicoabdominal artery. SNS (1 and 3 Hz) or injections of ACh (1.5 and 3 micrograms), DMPP (1 and 2 micrograms), and muscarine (1 and 2 micrograms) produced a frequency- or dose-dependent increase in both epinephrine (EPI) and norepinephrine (NE) output, determined from adrenal venous blood. Glucagon infusion (0.1, 0.3, and 1 microgram/min) enhanced the SNS-, ACh-, and DMPP-induced increases in EPI and NE output in a dose-dependent manner but did not affect the muscarine-induced increases in CA output. The increase in basal CA output induced by the highest dose of glucagon was only slight. Glucagon increased cyclic AMP overflow determined from adrenal venous blood. Our results indicate that glucagon has a facilitatory action on adrenal CA release mediated by nicotinic receptors but not by muscarinic receptors in dogs and suggest that an increase in cyclic AMP production in adrenal medullary cells may be responsible for its selective action.

Recently, soluble(s) circulating isoforms of intercellular adhesion molecule-1 (sICAM-1) and sE-selectin (formerly endothelial leukocyte adhesion molecule-1) have been described in normal human serum. Elevated levels have been reported in acute and chronic inflammatory disorders, including allograft rejection. In this study, plasma levels of sICAM-1 and sE-selectin were determined in groups of tacrolimus (FK 506)-treated adult patients following either isolated small bowel (SB), liver, or combined SB plus liver (SB/L) transplantation. Each molecule was measured at 1, 2, 4, 6, 8, and 12 weeks (all patients) and at 6, 9, and 12 months after transplantation (SB and SB/L only) by enzyme linked immunosorbent assay. Levels were compared with those of soluble interleukin-2 receptor (sIL-2R ; a marker of lymphocyte activation) and soluble HLA class I (which has been reported to be elevated in liver transplant-related complications). Elevations above normal in mean plasma levels of sICAM-1 (2.4-fold), sE-selectin (1.8-fold), sIL-2R (10.6-fold), and sHLA class I (1.3-fold) were found in patients with stable isolated SB grafts during the first 12 weeks posttransplant. Except for sHLA class I, levels of each protein were subsequently reduced, up to 1 year posttransplant. However, further increases in sICAM-1 and in sIL-2R and sE-selectin levels were observed during episodes of SB rejection compared with stable grafts. Mean levels of all molecules were higher in patients with isolated SB grafts compared with those given liver or combined (SB/L) transplants, either during stable SB graft function (up to 12 weeks posttransplant) or rejection. The data demonstrate increased adhesion molecule production/shedding following SB transplantation and are suggestive of a reduced overall level of immune activation in liver and SB/L compared with isolated SB transplantation.

We investigated the way in which hexamethonium and pirenzepine, a selective muscarinic (M) 1 receptor antagonist, modify the release of catecholamines from dog adrenal gland in response to endogenous and exogenous acetylcholine (ACh) in vivo. Output of epinephrine (EPI), and norepinephrine (NE) was determined from adrenal venous blood by high-performance liquid chromatography (HPLC) with electrochemical detection. Intraarterial (i.a.) injections of ACh (1.5 and 3 micrograms) or muscarine (3 and 6 micrograms) into the phrenicoabdominal artery and splanchnic nerve stimulation (SNS, 3 Hz) produced marked increases in EPI and NE output. Hexamethonium (1 mg/kg intravenously, i.v.) inhibited the increases in EPI and NE output in response to exogenous ACh and SNS. Pirenzepine (10 micrograms/kg i.v.) inhibited the ACh-induced increases in EPI and NE output but did not modify the SNS-induced increases. The muscarine-induced increases in EPI and NE output were also inhibited by pirenzepine. These results indicate that the exogenous ACh-induced release of adrenal catecholamines is mediated through both nicotinic and M1 receptors, in contrast to the predominant mediation of nicotinic receptors in response to endogenous ACh.

Effects of the renin-angiotension system (RAS) on adrenal catecholamine release in response to hemorrhagic hypotension and splanchnic nerve stimulation (SNS) were studied in pentobarbital-anesthetized dogs. In hemorrhage experiments, mean blood pressure (MBP) was maintained at 50 mm Hg for 60 min by bleeding the arterial blood into a pressurized bottle. In the renal intact group (control), epinephrine (EPI) and norepinephrine (NE) output from the adrenal gland increased markedly during hemorrhagic hypotension: from 45 +/- 13 and 4.7 +/- 0.9 to 1,167 +/- 202 and 169 +/- 30 ng/min at 60 min after onset of hemorrhage, respectively. The increases in catecholamine output during hemorrhagic hypotension in the renal-intact group pretreated with captopril (1 mg/kg intravenously, i.v.) and in the renal-ligated group were significantly smaller than those in the control group. The increases in catecholamine output in the renal-ligated group infused with angiotensin II (AngII 10 ng/kg/min i.v.) were comparable to those in the control group. In SNS experiments, AngII infusion (10 ng/kg/min i.v.) enhanced increases in catecholamine output induced by 3 Hz SNS significantly. Captopril (1 mg/kg i.v.) did not affect the SNS-induced increases in catecholamine output. These results suggest that the renal RAS facilitates reflex release of adrenal catecholamines during hemorrhagic hypotension, at least in part, by acting directly on the release process of catecholamines from dog adrenal gland.

Serotonin (5-HT 0.1 and 0.3 micrograms/kg) was injected as a bolus into the renal artery of anesthetized dogs. Renal blood flow (RBF) decreased initially after 5-HT injection and then increased. The 5-HT-induced decrease in RBF was potentiated during intrarenal infusion of a 5-HT1 and 5-HT2 antagonist methysergide at 3 micrograms/kg/min or of a 5-HT2 antagonist ketanserin (3-30 micrograms/kg/min), but methysergide at 30 micrograms/kg/min attenuated the decrease in RBF. The delayed increase in RBF was suppressed during ketanserin or methysergide infusion. In ketanserin-pretreated dogs, methysergide (3-30 micrograms/kg/min) dose-dependently suppressed the 5-HT-induced decrease in RBF. A 5-HT3 antagonist, ICS 205-930 (3-30 micrograms/kg/min), did not affect the 5-HT-induced RBF responses. A Ca2+ entry blocker CD-349 (30 and 100 ng/kg/min) and a Ca2+ release inhibitor TMB-8 (30 and 100 micrograms/kg/min) suppressed the 5-HT-induced decrease in RBF. These results suggest that 5-HT activates 5-HT1 receptors to induce vasoconstriction by mobilization of extracellular and intracellular Ca2+, but simultaneous activation of 5-HT2 receptors attenuates the vasoconstriction, probably by causing vasodilation in dog kidney.

The effects of dopamine (DA)-receptor and alpha 2-adrenoceptor agonists and antagonists on ganglionic muscarinic stimulation were examined in anesthetized dogs. All drugs were injected or infused intra-arterially into the blood supply of the cardiac sympathetic ganglia. The muscarinic agonists McN-A-343 (10, 20, and 30 micrograms) and muscarine (1, 2, and 3 micrograms) increased heart rate. The muscarinic receptor stimulation elicited by McN-A-343 or muscarine was significantly inhibited by infusion of the DA2-receptor agonist quinpirole (3 and 10 micrograms/min) but not by the DA1-receptor agonist SK&F 38393 (10 and 30 micrograms/min). The alpha 2-adrenoceptor agonist BHT 933 (3 and 10 micrograms/min) also inhibited muscarinic receptor stimulation. The DA2-receptor antagonist domperidone (10 micrograms/min) and the alpha 2-adrenoceptor antagonist yohimbine (1 micrograms/min) had no effects on muscarinic receptor stimulation, but they antagonized the inhibitory effects of quinpirole and BHT 933, respectively. The nicotinic transmission elicited by preganglionic cardiac sympathetic nerve stimulation (1 and 2 Hz) was also inhibited by DA-receptor and alpha 2-adrenoceptor agonists and antagonists. These results suggest that DA2-receptors and alpha 2-adrenoceptors suppress muscarinic transmission as well as nicotinic transmission in the cardiac sympathetic ganglia of the dog.

Effect of 3-4-dihydro-8-(2-hydroxy-3-isopropylaminopropoxy)-3-nitroxy-2H-1-b enzopyran (K-351) infused into the renal artery on renin release during graded renal nerve stimulation (RNS) was investigated in pentobarbital-anesthetized dogs. K-351 (20 micrograms/min) produced significant suppression of the RNS-induced renin release; the renin secretion responses to RNS at lower frequencies (0.5 and 1 Hz) were almost abolished, and those at the highest frequency (3 Hz) were attenuated. K-351 also suppressed an increase in renal vascular resistance during RNS at 3 Hz. The same extent of inhibition in the renin secretion response to RNS was also obtained during infusion of DL-propranolol (100 micrograms/min). Inhibitory effect of K-351, prazosin, or phentolamine on the renal vasoconstriction induced by RNS and norepinephrine (NE) injected into the renal artery, an approximately 50% reduction in renal blood flow, was also assessed. K-351 and prazosin exerted a greater inhibitory effect on RNS-than on NE-induced vasoconstriction, and the opposite was true of phentolamine. The potency of K-351 in reducing the vasoconstriction due to RNS or NE was roughly estimated to be 10-30 times less than that of prazosin. These results suggest that K-351 shares beta- and alpha-adrenoceptor blocking properties, which effectively contribute to the suppression of adrenergically induced renin release and renal vasoconstriction.

The effects of nifedipine, classed as a calcium entry blocker, on urinary excretion and renin release were investigated in anesthetized dogs. Nifedipine was infused into one renal artery for three consecutive 10-min periods, at incremental rates of 0.3, 1, and 3 micrograms/min. Intrarenal infusion of nifedipine (3 micrograms/min) produced marked increases in urine flow rate (by 200%) and in urinary sodium (by 210%) and potassium (by 40%) excretion rates of the infused kidney without changes in mean systemic blood pressure, heart rate, glomerular filtration rate, and filtration fraction. Urinary osmolarity was slightly decreased by the drug, but this change was not statistically significant. There were no consistent changes in these parameters for the contralateral noninfused kidney. Renin secretion rate was increased to three times the preinfusion level during infusion of the highest nifedipine dose. The blood flow to the infused kidney was increased, and ipsilateral renal vascular resistance was simultaneously decreased upon nifedipine infusion. These results suggest that a nonhypotensive dose of nifedipine causes an increase in renin secretion and that this drug has a striking effect on the reabsorption of sodium and water by the renal tubules.