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Abnormal vitamin A (retinol) metabolism plays an important role in the occurrence of non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH). In this study, NAFLD and NASH models were established to investigate the effects of food additives glycyrrhizic acid (GL) on retinol metabolism in NAFLD/NASH mice. Potential targets of GL and its active metabolite glycyrrhetinic acid (GA) were analyzed by RNA sequence, bioinformatics, and molecular docking analyses. Gene transfection and enzymatic kinetics were used to identify the target of GL. The results showed that GL could resolve the fatty and inflammatory lesions in the mouse liver, thereby improving the disorder of retinol metabolism. RNA sequence analysis of model mice liver revealed significant changes in AKR1B10 (retinol metabolic enzymes). Bioinformatics and molecular docking analyses showed that AKR1B10 is a potential target of GA but not GL. GA could inhibit AKR1B10 activity, which then affects retinol metabolism, whereas GL only had the same effect after hydrolysis into GA. In AKR1B10-KO hepatocytes, GA, GL, and hydrolysates of GL had no regulatory effect on retinol metabolism. Therefore, GA, the active metabolite of GL, as a novel AKR1B10 inhibitor, could promote retinoic acid synthesis. GL restored the balance of retinol metabolism in NAFLD/NASH mice by metabolizing to GA.

Non-steroidal anti-inflammatory drugs (NSAIDs)-induced small intestinal injury is a serious clinical event with recent advances of diagnostic technologies, but a successful therapeutic method to treat such injuries is still lacking. Licorice, a traditional herbal medicine, and its derivatives have been widely used for the treatment of a variety of diseases due to their extensive biological actions. However, it is unknown whether these derivatives have an effect on NSAIDs-induced small intestinal damage. Previously, the anti-inflammatory effects of three compounds extracted from the licorice root, glycyrrhizin, 18β-glycyrrhetinic acid, and dipotassium glycyrrhizinate, were compared in vitro cell culture. The most prominent inhibitory effect on the tumor necrosis factor-α (TNF-α) production was observed with the administration of 18β-glycyrrhetinic acid as an active metabolite of glycyrrhizin. In this study, a complex compound of 18β-glycyrrhetinic acid and hydroxypropyl γcyclodextrin was examined to improve the oral bioavailability. After administration of this complex to indomethacin treated mice, a significantly high plasma concentration of 18β-glycyrrhetinic acid was detected using the tandem mass spectrometry coupled with the HPLC. Furthermore, the complex form of 18β-glycyrrhetinic acid and hydroxypropyl γcyclodextrin reduced mRNA expressions of TNF-α, interleukin (IL)-1β, and IL-6, which was histologically confirmed in the improvement of indomethacin-induced small intestinal damage. These results suggest that the complex of 18β-glycyrrhetinic acid and hydroxypropyl γcyclodextrin has the potential therapeutic value for preventing the adverse effects of indomethacin-induced small intestinal injury.

([ 3 H]5-HT)-uptake and patch-clamp techniques were used to study the actions of (+) and (−) tramadol and the active metabolites of tramadol, (+) and (−) O-demethyl-tramadol on the human serotonin (5-HT) transporter and the human 5-HT 3A receptor, stably expressed in HEK-293 cells. The (+) and (−) enantiomers of tramadol suppressed the human 5-HT transporter concentration-dependently (IC 50 = 1.0 and 0.8 μM, respectively), resulting in 97% and 87% transport inhibition at their respective initial plasma concentrations (9.5 μM). The (+) and (−) enantiomers of the active tramadol metabolite were less potent than tramadol in inhibiting the human 5-HT transporter (IC 50 = 15 and 44 μM, respectively), resulting in 19.2% and 4.8% transport inhibition at their highest plasma concentrations (2.5 μM). In contrast to their potent suppression of the 5-HT transporter, both, (+) and (−) tramadol inhibited 5-HT (30 μM)-induced currents only at substantially higher concentrations (IC 50 = 199 and 251 μM, respectively), resulting in only 6% and 4% inhibition at the initial maximum plasma concentration. A similar low potent inhibition of human 5-HT 3A receptors was found for (+) and (−) O-demethyl-tramadol (IC 50 = 158 and 63 μM, respectively). In conclusion, at clinical plasma concentrations tramadol potently suppresses the human 5-HT transporter, whereas it has only a slight effect on the human 5-HT 3A receptor. The results are compatible with a possible mechanism for tramadol-induced early emesis involving the serotonergic system.

Nalmefene is an orally available opioid receptor antagonist that has been shown to suppress appetite in humans, but its effects on chronic food intake and body weight remain unclear. Here, we report that chronic (21-day) oral administration of nalmefene at 2 or 10 mg/kg/day in diet-induced obese (DIO) mice led to significant increases (9-11%) in cumulative food intake. Mice in the nalmefene-treated groups also gained body weight at a rate faster than the control. Body composition analysis showed that the extra body weight gains in the treated animals were mostly due to increased fat accumulation. Since acute nalmefene treatment showed a trend toward a decrease rather than an increase in food intake, it is possible that the orexigenic effect of chronic oral administration of nalmefene was caused by pharmacologically active metabolites rather than the drug itself. Our results argue against the potential use of nalmefene for treating human obesity.

The effects of tamoxifen, and its active metabolite endoxifen (4-hydroxy-N-desmethyl-tamoxifen), on hERG currents stably expressed in HEK cells were investigated using the whole-cell patch-clamp technique and an immunoblot assay. Tamoxifen and endoxifen inhibited hERG tail currents at -50mV in a concentration-dependent manner with IC50 values of 1.2 and 1.6μM, respectively. The steady-state activation curve of the hERG currents was shifted to the hyperpolarizing direction in the presence of endoxifen. The voltage-dependent inhibition of hERG currents by endoxifen increased steeply in the voltage range of channel activation. The inhibition by endoxifen displayed a shallow voltage dependence (δ=0.18) in the full activation voltage range. A fast application of endoxifen induced a reversible block of hERG tail currents during repolarization in a concentration-dependent manner, which suggested an interaction with the open state of the channel. Endoxifen also decreased the hERG current elicited by a 5s depolarizing pulse to +60mV to inactivate the hERG currents, suggesting an interaction with the activated (open and/or inactivated) states of the channels. Tamoxifen and endoxifen inhibited the hERG channel protein trafficking to the plasma membrane in a concentration-dependent manner with endoxifen being more potent than tamoxifen. These results indicated that tamoxifen and endoxifen inhibited the hERG current by direct channel blockage and by the disruption of channel trafficking to the plasma membrane in a concentration-dependent manner. A therapeutic concentration of endoxifen inhibited the hERG current by preferentially interacting with the activated (open and/or inactivated) states of the channel.

The molecular mechanism of action of dipyrone, a widely used antipyretic and non-opioid analgesic drug, is still not fully understood. Actions upon peripheral inflamed tissues as well as the central nervous system, especially upon the PAG-RVM axis, have been suggested. Dipyrone is a prodrug and its activity is due to its immediate conversion to its active metabolites. We tested the effect of two recently discovered metabolites of dipyrone, the arachidonoyl amides of 4-methylaminoantipyrine and 4-aminoantipyrine, on the neurons of the rostral ventromedial medulla (RVM), which are part of the descending pathway of antinociception. These compounds reduced the activity of ON-cells and increased the activity of OFF-cells. Both CB1 and TRPV1 blockade reversed these effects, suggesting that the endocannabinoid/endovanilloid system takes part in the analgesic effects of dipyrone.

We investigated the effects of rhein, an active metabolite of diacerein, on the interleukin-1alpha-stimulated production of nitric oxide (NO) in rabbit articular chondrocytes, and the effects of diacerein on NO production in rat adjuvant-induced arthritis. At doses of 10 and 30 microM, rhein significantly inhibited the interleukin-1alpha-stimulated NO production in chondrocytes. In the rat adjuvant-induced arthritis model, diacerein was administered for 21 days, starting at the time of adjuvant injection. Paw swelling and plasma NO level were measured in order to assess the effect of diacerein on arthritis and NO biosynthesis in the whole body. At doses of 30 and 100 mg/kg/day, diacerein significantly suppressed the development of adjuvant-induced arthritis and the increase in plasma NO. These results suggest that the inhibitory effect of diacerein on rat adjuvant-induced arthritis is partly related to its reduction of the NO production induced by adjuvant-induced arthritis.

The antimutagenic activity of eight novel imidazo[1,2-a]pyridine derivatives (I-VIII) against sodium azide (NaN3) and benzo[a]pyrene (B[a]P) was evaluated using the Salmonella reverse mutation assay. At non-toxic concentrations (12.5–50 µM), imidazopyridines I, II, III, and V with a terminal imidazopyridine group were mutagenic, while derivatives VII and VIII with a central imidazopyridine group were not mutagenic. Compounds IV, VII, and VIII exerted a moderate antimutagenic activity against NaN3 under pre-exposure conditions, and a strong activity (>40%) against B[a]P in the presence of S9 under both pre- and co-exposure conditions and mostly independent on the dose. Imidazopyridines possibly inhibited the microsomal-dependent activation of B[a]P. The demethylated derivative VII was the most active antimutagen. All imidazopyridines had a low to moderate antioxidant activity. The antibacterial activity of imidazopyridines was sporadic and moderate probably due to the failure of bacteria to convert imidazopyridines into active metabolites. The position of imidazopyridine was a pivotal factor in the mutagenic/antimutagenic activity. The strong antimutagenic compounds were dicationic planar compounds with a centered imidazo[1,2-a]pyridine spacer. With LD50 of 60 mg/kg in mice for both derivatives VII and VIII, it is safe to investigate the anticancer activity of these derivatives in animal models.

Recent studies have reported elevated expression of cluster of differentiation (CD) 147 on CD14(+) monocytes of the peripheral blood of patients with active rheumatoid arthritis and a correlation of CD147 expression with Disease Activity Score. Thus, CD147 may be a new target for treatment of rheumatoid arthritis. Leflunomide is a disease-modifying antirheumatic drug that is commonly used to treat rheumatoid arthritis. The effect of leflunomide in blocking the up-regulation of CD147 and in blocking the down-regulation of metalloproteinases (MMP)-2 and MMP-9 in active macrophages has not yet been established. In this study we investigated the effect of A771726, the active metabolite of leflunomide, on expression of CD147 and on the gelatinolytic activity of MMP-2 and MMP-9 in phorbol myristate acetate (PMA) differentiated THP-1 cells. The expression of CD147, MMP-2, and MMP-9 mRNAs were determined by real-time quantitative reverse transcription PCR, the levels of cellular surface expression of CD147 were determined by flow cytometry, and the gelatinolytic activity of MMP-2 and MMP-9 were determined by zymography. Our results showed that A771726 significantly inhibited the expression of CD147 on the cell surface of activated THP-1 cells in a dose-dependent manner (P

The importance of d-norfenfluramine in regard to the indole-depleting action of d-fenfluramine has not been well studied in sensitive animal species. The present study therefore examined the intensity and time course of the neurochemical effects of i.p. injected d-fenfluramine (2.5 and 5 mg/kg) and d-norfenfluramine (2.5 mg/kg) in vehicle- and SKF-525A-pretreated rats, relating the effects to the brain concentration-time profiles of the drug and its active metabolite. At the lower dose d-fenfluramine caused only a small, short-lasting decrease in brain serotonin (5-HT) without affecting the 5-hydroxyindoleacetic acid (5-HIAA). Higher doses affected both 5-HT and 5-HIAA (50-60 and 30-40% reductions, respectively), the effect being maximal for at least 8 h. d-Norfenfluramine reduced the brain content of 5-HT and 5-HIAA less (by about 30%) than 5 mg/kg d-fenfluramine did. Brain concentrations of d-norfenfluramine at the time of the maximal depletion of indoles were close to those of the metabolite after 5 mg/kg d-fenfluramine, indicating that the acute indole-depleting effects did not depend solely on the brain concentrations of its nor-metabolite. SKF-525A changed the metabolite-to-parent drug ratios in brain without appreciably influencing the action of d-fenfluramine. However, the maximum decrease in indole content caused by 2.5 mg/kg d-fenfluramine in SKF-525A-pretreated rats was only 12% of the control level, although the brain concentration of unchanged drug was comparable to that after 5 mg/kg d-fenfluramine in vehicle-pretreated rats.(ABSTRACT TRUNCATED AT 250 WORDS)

The pharmacological profile of a novel angiotensin II type 1 receptor blocker, azilsartan medoxomil, was compared with that of the potent angiotensin II receptor blocker olmesartan medoxomil. Azilsartan, the active metabolite of azilsartan medoxomil, inhibited the binding of [(125)I]-Sar(1)-I1e(8)-angiotensin II to angiotensin II type 1 receptors. Azilsartan medoxomil inhibited angiotensin II-induced pressor responses in rats, and its inhibitory effects lasted 24h after oral administration. The inhibitory effects of olmesartan medoxomil disappeared within 24h. ID(50) values were 0.12 and 0.55 mg/kg for azilsartan medoxomil and olmesartan medoxomil, respectively. In conscious spontaneously hypertensive rats (SHRs), oral administration of 0.1-1mg/kg azilsartan medoxomil significantly reduced blood pressure at all doses even 24h after dosing. Oral administration of 0.1-3mg/kg olmesartan medoxomil also reduced blood pressure; however, only the two highest doses significantly reduced blood pressure 24h after dosing. ED(25) values were 0.41 and 1.3mg/kg for azilsartan medoxomil and olmesartan medoxomil, respectively. In renal hypertensive dogs, oral administration of 0.1-1mg/kg azilsartan medoxomil reduced blood pressure more potently and persistently than that of 0.3-3mg/kg olmesartan medoxomil. In a 2-week study in SHRs, azilsartan medoxomil showed more stable antihypertensive effects than olmesartan medoxomil and improved the glucose infusion rate, an indicator of insulin sensitivity, more potently (≥ 10 times) than olmesartan medoxomil. Azilsartan medoxomil also exerted more potent antiproteinuric effects than olmesartan medoxomil in Wistar fatty rats. These results suggest that azilsartan medoxomil is a potent angiotensin II receptor blocker that has an attractive pharmacological profile as an antihypertensive agent.

The neuromuscular-blocking agent vecuronium bromide undergoes hydrolysis to three pharmacologically active metabolites (3-desacetyl, 17-desacetyl and 3,17-desacetyl vecuronium) which might modify the neuromuscular-blocking action of their parent compound. In order to elucidate the possible role of the interaction between vecuronium and its metabolites in the complications reported after long-term use of vecuronium in intensive care unit (ICU) patients, the relative potency of vecuronium, 3-desacetyl and 3,17-desacetyl vecuronium was determined in the rat hemidiaphragm in vitro and the mode of interaction of the above-mentioned compounds investigated. Dose-response relationships were established for each substance alone and for combinations of vecuronium with its metabolites. The relative potency at the EDmax50 levels (% maximal effect) were in the order of 1:1.2:27 for vecuronium, the 3-desacetyl derivative and the 3,17-desacetyl derivative, respectively. The mode of interaction characterized by isobolographic and algebraic (functional) analysis showed vecuronium and 3-desacetyl vecuronium to interact in an additive fashion while the combined effect of the parent compound and its 3,17-desacetyl derivative was less than additive, indicating antagonism.

The present study was undertaken to investigate the role of prostaglandins in the shortening of transit time observed after intraduodenal administration of rhein anthrone and rhein. After intraduodenal administration of rhein anthrone (0.5-10 mg/rat), a dose-dependent acceleration of small intestinal transit was observed. The effect for rhein (1-10 mg/rat) was far less pronounced. In the same test conditions, analysis of small intestinal tissue revealed a significant increase of prostaglandin E2 (PGE2), reaching its maximum value 30 min after administration of rhein anthrone. The increase in PGE2 found 30 min after administration of rhein was not significant. The effects provoked by rhein anthrone could be largely prevented by pretreatment of the animals with indomethacin (1-3 mg rat) or cortisol (10 mg/rat). It is concluded that prostaglandins play an important role in the acceleration of the transit provoked in rats by rhein anthrone.

Neuropsychiatric adverse events have been reported in influenza patients with and without exposure to oseltamivir (Tamiflu®), triggering speculation as to whether oseltamivir may be interacting with any human receptors and contributing to such neuropsychiatric events. In this study, the in vitro selectivity profile of oseltamivir prodrug and active metabolite was investigated. Both compounds lacked clinically relevant pharmacological activities on human, rodent and primate neuraminidases and on a panel of 155 other molecular targets, including those relevant for mood, cognition and behavior. Neuropsychiatric adverse events observed in influenza patients are likely a phenomenon caused by the infection rather than by oseltamivir.

Interpretation of the in vivo actions of 3,4-methylenedioxymethamphetamine (MDMA) is complicated by the formation of the active metabolite, 3,4-methylenedioxyamphetamine (MDA). This study evaluates the role of MDA in the dopamine releasing actions of (+)-MDMA. In the study, rats were given subcutaneous doses of (+)-MDMA and concentrations of monoamines and their metabolites in striatal dialysate were measured at 15 min intervals. In parallel experiments, plasma concentrations of (+)- and (-)-MDMA and MDA were determined by GC/MS procedures. The time course of MDMA levels was comparable for the two isomers as were their bioavailabilities. In contrast, the plasma levels of MDA were about three times higher after (+)-MDMA. (+)-MDMA caused a rapid increase in striatal dialysate levels of dopamine and decreased extracellular levels of dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). There was a significant correlation between dopamine concentration in striatal dialysate and plasma MDMA concentration, but not with plasma MDA. These results indicate that MDMA itself has stereoselective actions on dopamine neurons. However, the higher plasma MDA levels after (+)-MDMA may account for part of the enantiomeric differences in the behavioral and neurotoxicological effects of MDMA.

One of the major factors limiting the use of non-steroidal anti-inflammatory drugs is gastrointestinal toxicity. Gaultherin, 2-[(6-O-beta-D-Xylopyranosyl-beta-D-glucopyranosyl)oxy] benzoic acid methyl ester, a natural salicylate derivative extracted from Gaultheria yunnanensis, has been shown to have analgesic and anti-inflammatory effects and lack gastric ulcerogenic effect compared to aspirin in our primary study. The aim of this study was to investigate the mechanism of action of gaultherin, which may rely on its active metabolite, and the mechanism responsible for the non-ulcerogenic property. The results showed that gaultherin (200 mg/kg) significantly inhibited the abdominal contractions in the acetic acid-induced writhing test in mice. The anti-inflammatory effect of gaultherin was demonstrated in the croton oil-induced ear edema model in mice. The results showed that gaultherin and equimolar dose of aspirin produced comparable inhibitory effects. The study of the metabolism characters of gaultherin in mice and rats indicated that gaultherin could be metabolically converted to salicylate, which produced the pharmacological effects, and provided effective concentrations for an extended period. In vitro metabolism experiment showed that gaultherin was metabolized by beta-glycosidase produced by human intestinal bacteria and esterases in intestine, blood and liver successively to release salicylate finally. The study suggested gaultherin did not cause gastric ulcer for the reason that it released salicylate in intestine slowly, not in stomach and it left the cyclooxygenase-1 unaffected, which was the source of cytoprotective prostaglandins in gastric epithelium.

K579 ((S)-1-[4-methyl-1-(2-pyrimidinyl)-4-piperidylamino]acetyl-2-pyrrolidinecarbonitrile), which is a long-acting and a slow binding dipeptidyl peptidase IV inhibitor, preserved the endogenously secreted active forms of glucagon-like peptide-1, augmented the insulin response and ameliorated the glucose excursion during oral glucose tolerance test in rats. In this study, we measured plasma concentrations of K579 after oral administration to rats. However, K579 was eliminated rapidly from plasma after oral administration to rats. Therefore, we postulated that there are active metabolites of K579 in rat plasma. We investigated the effect of K579 on plasma dipeptidyl peptidase IV activity using bile duct-cannulated rats. The duration of inhibitory action of plasma dipeptidyl peptidase IV after the administration of K579 in bile duct-cannulated rats was shorter than that in sham-operated rats. Moreover, we investigated the effect of bile obtained from K579-treated rat on plasma dipeptidyl peptidase IV activity in normal rats. The bile collected from K579-treated rats exhibited tardive and potent inhibitory activity of normal rat plasma. These results suggest that K579 sustained the duration of inhibitory action of plasma dipeptidyl peptidase IV by the character as a slow-binding inhibitor and, as well, by the presence of metabolites of K579, which exhibit the inhibitory activity of dipeptidyl peptidase IV.

The concentration dependences of the Ca(2+)-sensitizing and the phosphodiesterase-inhibitory effects of levosimendan (the (-) enantiomer of [[4-(1,4,5,6-tetrahydro-4-methyl-6-oxo-3-pyridazinyl)phenyl]hydrazono]propanedinitrile) and its active metabolite, OR-1896 (the (-) enantiomer of N-[4-(1,4,5,6-tetrahydro-4-methyl-6-oxo-3-pyridazinyl)phenyl] acetamide), were compared with their positive inotropic effects to reveal their mechanisms of action in guinea pig hearts. In Langendorff-perfused hearts, left ventricular +dP/dt(max) increased by 26+/-4% and 25+/-3% (mean+/-S.E.M.), with EC(50) values of 15+/-2 and 25+/-1 nM for levosimendan and OR-1896, respectively. In permeabilized myocyte-sized preparations, levosimendan and OR-1896 both increased isometric force production via Ca(2+) sensitization (at pCa 6.2), by 51+/-7% and 52+/-6%, with EC(50) values of 8+/-1 and 36+/-7 nM (P

Prulifloxacin, a new thiazeto-quinoline derivative with antibiotic properties, was evaluated for cardiac risk both in vitro on the ether-à-go-go-related gene (HERG) K+ channel, and in vivo in the conscious dog monitored by telemetry. HERG current was measured from stably transfected human embryonic kidney (HEK) 293 cells by means of the patch-clamp technique. Application of AF 3013, the active metabolite of prulifloxacin, produced only minor reduction of HERG current amplitude (tail current=-40 mV), producing a maximum blockade of 12.3 +/- 3.3% at the highest concentration tested (335 microM). In comparison, ciprofloxacin also failed to produce a 50% inhibition of HERG current amplitude, although the maximum blockade was greater than that observed with prulifloxacin (47.6 +/- 1.9% at the highest concentration tested (335 microM). In contrast, moxifloxacin blocked HERG current amplitude with an IC50 value of 74.7 microM. Prulifloxacin had no effect on the QTc interval (Fridericia's) following 5 days of repeated oral administration (150 mg/kg/day) in the conscious dog monitored by telemetry. These findings suggest that prulifloxacin is not likely to prolong the QT interval.

l-alpha-acetylmethadol is a long-acting narcotic analgesic that is used in the treatment of opiate addiction. However, the drug has been associated with cases of QT interval prolongation and ventricular arrhythmia. To understand the mechanism underlying these clinical findings, we examined the effects of l-alpha-acetylmethadol on the cloned human cardiac K(+) channels HERG (human ether-a-go-go-related gene), KvLQT1/minK and Kv4.3. Using patch clamp electrophysiology, we found that l-alpha-acetylmethadol inhibited HERG channel currents in a voltage-dependent manner displaying an IC(50) value of 3 microM. The major active metabolite of l-alpha-acetylmethadol, noracetylmethadol, inhibited HERG with an estimated IC(50) values of 12 microM. l-alpha-acetylmethadol had little or no effect on Kv4.3 or KvLQT1/minK K(+) channel currents at concentration up to 10 microM. We conclude that the proarrhythmic effects of l-alpha-acetylmethadol are due to specific blockade of the HERG cardiac K(+) channel and that its active metabolite noracetylmethadol may provide a safer alternative in the treatment of opiate addiction.

We performed experiments in dog ventricular trabeculae loaded with aequorin to elucidate the mechanism of positive inotropic effect of (R)-N-[4-(4-methyl-6-oxo-1,4,5, 6-tetrahydro-pyridazin-3-yl)-phenyl]-acetamide (OR-1896), an active metabolite of (R)-([4-(1,4,5, 6-tetrahydro-4-methyl-6-oxo-3-pyridazinyl)phenyl]-hydrazono)-pr opaned initrile (levosimendan). Concentration-response curve for OR-1896 was biphasic: positive inotropic effect of OR-1896 reached a plateau at 10(-5) M (1st phase) and the concentration-response curve became steeper at 10(-3) M and higher (2nd phase). Maximum response of the 1st phase was 29% of maximal response to isoproterenol and associated with an increase in Ca(2+) transients of 13% of the maximal response to isoproterenol. For a given increase in force, the increase in Ca(2+) transients by OR-1896 was lower than that induced by elevation of [Ca(2+)](o). The positive inotropic effect of OR-1896 was not associated with impairment of relaxation and it was abolished by carbachol. In conclusion, OR-1896 has a positive inotropic effect partly due to an increase in myofibrillar Ca(2+) sensitivity that is exerted via cross-talk with signal transduction mediated by cAMP.

The antithrombotic effect of desethyl KBT-3022, which is the main active metabolite of the new antiplatelet agent, KBT-3022 (ethyl 2-[4,5-bis(4-methoxyphenyl)thiazol-2-yl] pyrrol-1-ylacetate; a cyclooxygenase inhibitor), was determined using a photochemically induced arterial thrombosis model in the rat femoral artery. Pretreatment with desethyl KBT-3022 (0.1, 0.3 and 1 mg/kg, i.v.) prolonged the time required to achieve thrombotic occlusion in the femoral artery and inhibited collagen-induced platelet aggregation in whole blood ex vivo, each in a dose-dependent manner. In all 6 rats used, particularly at the highest dose (1 mg/kg, i.v.) tested, cyclic variations in blood flow were hardly ever observed and complete cessation of blood flow did not occur during the 30-min observation time. BM-13505 (1, 3 and 10 mg/kg, i.v.), a thromboxane A2 receptor antagonist, also prolonged the time to occlusion, but cyclic variations in blood flow did occur. On the other hand, aspirin (10 and 30 mg/kg, i.v.) had little effect in terms of preventing thrombosis, although it inhibited collagen-induced platelet aggregation to the same extent as did desethyl KBT-3022. Desethyl KBT-3022 inhibited the thrombin-induced aggregation of washed platelets in a concentration-dependent manner (1–40 μM), whereas aspirin and BM-13505 did not. These findings suggest that the potent antithrombotic effect of desethyl KBT-3022 may be attributable in part to its additional ability to inhibit thrombin-induced platelet aggregation. Accordingly, thromboxane A2 and thrombin may be important thrombotic mediators in this rat model.

(+/-)-1-(Cyclohexyloxycarbonyloxy)ethyl 2-ethoxy-1-[[2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl]-1H- benzimidazole-7-carboxylate (TCV-116, Candesartan) and its active metabolite 2-ethoxy-1-[[2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl]-1H- benzimidazole-7-carboxylic acid (CV-11974) are specific nonpeptide angiotensin AT1 receptor antagonists. In the present study, the inhibitory potency of these two antagonists on the angiotensin II-induced responses in aortic vascular smooth muscle cells from Wystar Kyoto rats was investigated. The specific binding of 125I-angiotensin II to cells was inhibited by CV-11974 and TCV-116 with a half-maximal inhibitory concentration (IC50) of 3 x 10(-11) M and 1 x 10(-9) M, respectively. CV-11974 and TCV-116 inhibited the angiotensin II-induced increase in [3H]thymidine incorporation with an IC50 of 3 x 10(-10) and 5 x 10(-9) M, respectively. Both CV-11974 and TCV-116 (10(-7) M) completely blocked the angiotensin II-induced increase in c-fos mRNA. The inhibitory potency of the metabolite CV-11974 was about 30-100-fold higher than that of the prodrug TCV-116.

The purpose of this study was to determine whether there are age-related changes in the extent of in vitro-induced nitrate tolerance. Nitroglycerin pre-exposure (10 μM for 30 min) provoked a significant shift to the right of the dose-response curve to nitroglycerin in aortae isolated from rats of 8 weeks, 12 and 18 months. However, this shift to the right was significantly larger at 18 months, both when KCl or phenylephrine were used as contractile agents. On the contrary, nitroglycerin pre-exposure did not significantly alter the dose-dependent relaxation to Sin-1 (3-morpholinosydnonimine, the active metabolite of molsidomine) at 8 weeks, 12 and 18 months. These data indicate that the extent of the in vitro-induced nitrate tolerance is larger when aortae are isolated from senescent rats. This increase in tolerance does not appear to involve desensitization of guanylate cyclase.

The Ca2+ entry blocking effects of MPC-1304, a new Ca2+ entry blocker of the 1,4-dihydropyridine type, and of its (S) and (R) enantiomers and metabolites were examined on Ca2+-induced contractions in isolated rabbit arteries. The Ca2+ entry blocking activity of the (S) enantiomer of MPC-1304 was approximately 150 times greater that that of its (R) enantiomer. Likewise, the antihypertensive effect of the (S) enantiomer was twice as great as that of MPC-1304 (racemate) in conscious spontaneously hypertensive rats, while the (R) enantiomer was ineffective. Thus, most of the pharmacological activity of MPC-1304 resides in its (S) configuration. The main metabolic products of MPC-1304 lso inhibited the Ca2+-induced contraction in the isolated vascular smooth muscles. These active metabolites showed a stereoselectivity similar to that of MPC-1304 for Ca2+ entry blocking activity, and may contribute to the potent antihypertensive action of MPC-1304.

The diuretic activity of a quinolinone oxime diuretic, M12285, was examined after renal arterial, i.v. and portal injection in rats. M12285 injected into the renal artery at a dose of 1 mg/kg caused no diuretic effect, whereas i.v. and portal injections induced marked diuresis dose dependently. The minimum effective dose with portal injection was lower (1 mg/kg) than that with i.v. injection (3 mg/kg) and the start of the effect was faster with portal injection. These results indicated that some metabolic modification in the liver is necessary for the diuretic activity to appear. Accordingly, we performed in situ rat liver perfusion with M12285 and obtained several metabolites. Renal arterial injection of each fractionated metabolite of M12285 revealed that all the diuretic activity derived from one of these metabolites. From IR and 1H-nuclear magnetic resonance (1HNMR) measurements, the chemical structure of this active metabolite was assumed to be a sulfate-conjugated form of M12285 at the oxime moiety. Based on this tentative chemical structure, we synthesized the oxime sulfate of M12285 (potassium salt, M17000) and confirmed the identity of IR and 1HNMR spectra. Administration of M17000 into the renal artery induced apparent diuresis in a dose-dependent manner in both rats and dogs. These results indicate that the oxime sulfate of M12285 is responsible for the diuretic activity of M12285. Therefore, we synthesized several derivatives of M17000 and confirmed their possible therapeutic value as a novel family of diuretics, namely quinolinone oxime sulfonic acids.

In contrast with furosemide (and other sulfamoylbenzoic and aryloxyacetic acids loop diuretics), MK-447 was unable to inhibit the [Na+,K+,Cl-] cotransport system in human red blood cells. Indeed, this compound was a very poor ion transport inhibitor (inactive on Ca(2+)-sensitive K+ channels, the Ca2+ pump, the Na+:Mg2+ exchange, the Na+:Li+ countertransport system and the [K+,Cl-] cotransport system, and only inhibiting the [Cl-/HCO3-] exchanger and the Na+,K+ pump at high concentrations). Conversely, its urinary metabolite (O-sulfo)-MK-447 was a very potent inhibitor of the [Na+,K+,Cl-] cotransport system (IC50 of 1.6 +/- 0.5 x 10(-6) M; mean +/- S.D. of four experiments). This compound was a much more potent [Na+,K+,Cl-] cotransport inhibitor than furosemide, and almost as active as bumetanide. In addition, (O-sulfo)-MK-447 was a moderate inhibitor of the [Cl-/HCO3-] exchanger (IC50 of 6 +/- 3 x 10(-5) M, n = 3), its potency being intermediate between that of xipamide and that of furosemide. Interestingly, it exhibited some inhibitory activity against Ca(2+)-sensitive K+ channels but only at high concentrations (it had no effect on the [K+,Cl-] cotransport system, the Ca2+ pump or the Na+:Mg2+ exchanger). The results suggest strongly that the O-sulfo derivative of MK-447 is an active natriuretic metabolite of MK-447. This metabolite may be responsible for the salidiuretic action of MK-447.

The effects of vitamin D3 and two analogues on embryonic angiogenesis were studied in 4.5-day-old chick embryo chorioallantoic membranes. The active metabolite of vitamin D3, 1 alpha,25-dihydroxyvitamin D3, and a synthetic vitamin D3 analogue, 22-oxa-1 alpha,25-dihydroxyvitamin D3, inhibited angiogenesis in a dose-dependent manner, the inhibition occurring in the picomolar range. In contrast, vitamin D3 was not effective. The results suggest that these two vitamin D3 analogues might be promising anti-angiogenic agents for controlling the angiogenesis which occurs in several pathological conditions, including tumor development.

The effect of L-cysteine on the depletion of serotonin and 5-hydroxyindoleacetic acid concentrations caused by p-chloroamphetamine and d-fenfluramine was studied in various brain regions one week after drug injection. p-Chloroamphetamine (2.5 and 5 mg/kg i.p.) and d-fenfluramine (13.4 mg/kg i.p.) significantly reduced serotonin and 5-hydroxyindoleacetic acid levels in the striatum, hippocampus and cortex, particularly in the latter areas. L-cysteine (500 mg/kg i.p.), administered 30 min before and 5 h after p-chloroamphetamine or d-fenfluramine, significantly reduced the effect of either drug on the concentrations of both indoles without causing any effect by itself. In another experiment, the rats were treated as above and were killed at various times after p-chloroamphetamine or d-fenfluramine injection to determine, in parallel, the indole levels in the whole brain and concentration of p-chloroamphetamine, d-fenfluramine and its metabolite d-norfenfluramine in the plasma and brain. p-Chloroamphetamine and d-fenfluramine markedly lowered both indoles, particularly 16 and 24 h after injection. L-cysteine had no effect on the indole concentrations but significantly reduced the effect of p-chloroamphetamine and d-fenfluramine 16 and 24 h after injection. At these times, the brain concentrations of p-chloroamphetamine, d-fenfluramine and d-norfenfluramine were markedly lower in the L-cysteine-treated than in the control rats. Analysis of the blood concentration of p-chloroamphetamine, d-fenfluramine and d-norfenfluramine showed that the rats treated with L-cysteine eliminated the drugs studied more rapidly than the control animals. The results show that L-cysteine partially prevents the depletion of the brain content of serotonin and 5-hydroxyindoleacetic acid caused by p-chloroamphetamine and d-fenfluramine one week after drug administration. The fact that the effect of p-chloroamphetamine and d-fenfluramine seen at shorter times was associated with a marked reduction in the blood and brain drug concentration in the L-cysteine-treated animals suggests that the reduced availability of p-chloroamphetamine, d-fenfluramine and its active metabolite may be responsible for the partial prevention by L-cysteine of their neurochemical effects.

Piribedil (ET495) and its catechol metabolite S584 induced enhanced stereotyped behaviour and locomotor activity in adult rats treated as neonates with intraventricular 6-hydroxydopamine. These same rats also showed an enhanced behavioural response to apomorphine, a directly acting dopamine agonist, but a blocked behavioural response to amphetamine, an indirectly acting dopamine agonist. This suggests that the dopaminergic action of piribedil is postsynaptic in origin and may be mediated via its active metabolite S584.

The effect of acid and non-acid antipyretic analgesics on prostaglandin (PG) release from cultured mouse astrocytes and peritoneal macrophages was investigated in order to test the hypothesis that the non-acid compounds are more potent inhibitors of PG formation in brain than in peripheral tissues. Stimulation of the cells by the divalent cation ionophore A 23187 (10 −6 mol/1) induced PG release from astrocytes and macrophages (mainly PGD 2 and PGE 2 , respectively). This PG release was inhibited by acetylsalicylic acid (10 −5 – 10 −6 mol/1) and indomethacin (10 −6 – 10 −9 mol/1) but also by high concentrations (10 −3 – 10 −5 mol/1) of the non-acid compounds 4-methyl-aminophenazone, the main active metabolite of dipyrone (metamizol), and acetaminophen (paracetamol). No difference was found in the inhibitory potency of the drugs in astrocytes and macrophages, suggesting that a specific sensitivity of brain cells toward non-acid antipyretic analgesics does not contribute to their analgesic effect.

The i.p. LD 5 0 of the organophosphorus ester triamiphos to adult male rat is about 15 mg/kg. The symptoms are typical of peripheral cholinesterase inhibition. Thus death is due to respiratory paralysis; there is no stimulation of the central nervous system. Measurement of the cholinesterase activity of several tissues after i.p. administration of various doses of triamiphos indicates that: (1) the rapidity of true cholinesterase inhibition at the myoneural junction is a major factor in the outcome of the acute intoxication; (2) brain true cholinesterase is not significantly inhibited; (3) in vivo, pseudocholinesterase is more inhibited than true cholinesterase; (4) except in the intestine, no return of cholinesterase activity was observed within 18 hr following the injection. Triamiphos is not a direct cholinesterase inhibitor but conversion by the liver to an anticholinesterase agent has been demonstrated by incubation of triamiphos with a fortified liver preparation. The active metabolite is very labile and does not readily cross the blood brain-barrier.

The anticonvulsant activity of valproic acid (VPA) was determined in amygdala-kindled rats after single and repeated (total of 7 injections given 3 times per day) administration of 200 mg/kg i.p. After a single injection, VPA significantly reduced the severity and duration of the kindled seizures and increased the duration of after-discharges recorded from the stimulated amygdala, but only 12% of the animals were totally protected from seizures. The percentage of animals totally protected increased to 88% after 7 doses. This pronounced increase in the anticonvulsant activity was not related to alterations in the plasma concentrations of VPA or its major active metabolites. Furthermore, determination of VPA and its metabolites in the substantia nigra after a single and repeated administration yielded the same data, again indicating that the increase in the anticonvulsant activity was not due to drug accumulation. In contrast to the marked increase in the anticonvulsant efficacy of VPA during short-term treatment, wet-dog shake behaviour induced by a single injection of the drug was significantly attenuated after repeated dosing, indicating that the anticonvulsant effect of VPA and the wet-dog shake behaviour induced by the drug were not mediated by the same mechanism. This was substantiated by experiments with one of the major metabolites of VPA in rat plasma, trans-2-en-VPA, which had approximately the same anticonvulsant efficacy as VPA but did not induce wet-dog shakes in rats.

A comparative study of the ability to block the amine pump was carried out on tricyclic antidepressants including dothiepin and northiaden in vivo. Dothiepin was found to prevent the 6-OHDA-induced depletion of cardiac noradrenaline but not the PCA-induced depletion of intracerebral serotonin. Northiaden, an active metabolite of dothiepin, also possessed the same ability with a greater potency than the parent drug. Neither compound affected the biosynthesis of catecholamines and indoleamine. However, neither dothiepin nor northiaden affected 5-HT uptake, as was also observed with imipramine and amitriptyline. These results suggest that the clinical efficacy of dothiepin may be due to inhibition of the amine pump, especially of the catecholamine uptake mechanism, which is qualitatively the same as for imipramine and amitriptyline.

Rats were injected with chlorpromazine (CPZ), 21 μmoles/kg (7.5 mg/kg) i.p., and killed after different time intervals up to 24 h. Mass fragmentographic methods were used to determine the levels of CPZ, monodemethyl-chlorpromazine (nor1-CPZ) and 7-hydroxy-chlorpromazine (7-OH-CPZ) in brain and blood and dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxy-3-indoleacetic acid (5-HIAA) in the straitum. Significant correlations between the brain and blood concentrations of CPZ as well as its active metabolites and the levels of DOPAC and HVA were obtained at several time points. The similarity between the time curves for CPZ — but not its metabolites — and the elevation of the dopamine (DA) metabolites strongly suggest that it is predominantly the unchanged drug in the brain which determines the acceleration of DA metabolism.

A recently discovered metabolite, diethylthiocarbamic acid methyl ester (Me-DTC), has been found in the plasma of man and rats in much higher concentrations than any other described metabolite after therapeutic doses of disulfiram. Me-DTC, in contrast to other disulfiram metabolites, is a potent inhibitor of liver aldehyde dehydrogenase (ALDH) in vitro. Like disulfiram, Me-DTC had a pronounced hypothermic effect in rats. This hypothermic effect and the augmented blood pressure response to ethanol challenge in rats developed rapidly with Me-DTC but were somewhat delayed with disulfiram. The blood pressure response outlasted the presence of Me-DTC in plasma (less than 24 h); a significant effect was found 48 h after pretreatment but not 72 h after a single dose. No effect was observed when ethanol was given 15 min before Me-DTC or disulfiram. These latter two observations are consistent with a function of Me-DTC as a suicide inhibitor of ALDH. Since Me-DTC has been reported to inhibit ALDH in vitro, even under anaerobic conditions, Me-DTC may be the active metabolite of disulfiram.

The anticonvulsant activity of propranolol and two selected metabolites, propranolol glycol and N-desisopropylpropranolol were compared in mice against 4 types of experimentally induced seizures: pentylenetetrazol, strychnine, low frequency and maximal electroshock. Both metabolites possessed significant anticonvulsant activity with propranolol glycol being 1/2 to 1/3 as potent and N-desisopropylpropranolol being 1/6 as potent as propranolol. The possible contribution of these two active metabolites to the acute anticonvulsant efficacy of propranolol was assessed in time course studies. Maximal anticonvulsant activity occurred between 2.5--10 min after propranolol (5-20 mg/kg, i.v.) and was significantly diminished after 30 min. Following propranolol, brain levels of both metabolites were extremely low ( less than 10 ng/g) at the onset of anticonvulsant action and reached peak levels between 15-30 min at which time anticonvulsant activity was already declining. In contrast, brain levels of propranolol were similar in time course to that observed for its anticonvulsant effect and there was significant positive correlation between these two parameters in 3 of the 4 seizure models. These data indicate that although these two metabolites are pharmacologically active, they do not contribute significantly to the acute anticonvulsant actions observed after propranolol administration.

In vitro receptor binding studies with paraoxon treated rat brain membranes, and stability studies using HPLC analysis, showed a negligible opiate receptor affinity for the morphinomimetic agent nicomorphine and its hydrolysis product 3-nicotinoylmorphine. Two other hydrolysis products, 6-nicotinoylmorphine and morphine, had a high affinity in the nanomolar range and are probably the active metabolites of nicomorphine.

There is a lag phase of 30-60 min before the onset of bromocriptine (BC) action. This delay may be necessary for the formation of active metabolites. The objective was to determine whether the abnormal behavioral effects induced by BC involve active hepatic metabolites. Thus, we studied the effect of an inhibitor of hepatic hydroxylation metabolism (SKF 525A) on the behavior of BC-treated cats. Experiments began after six weeks of habituation and involved i.p. injections of: (1) propylene glycol (drug vehicle); (2) SKF 525A (70 mg/kg); (3) BC (10 mg/kg); and (4) SKF 525A followed 30 min later by BC. Each cat received the four treatments with two weeks elapsing between consecutive experiments. The frequency of 12 behaviors was scored for 60 min after 1 h posttreatment. BC alone induced emergent behavioral changes (hallucinatory-like, limb flicks, abortive grooms) that were not observed following control injections (vehicle and SKF 525A). There was a complete elimination of BC-induced hallucinatory-like behavior/escape by SKF 525A pretreatment. Other emergent behaviors were similarly reduced but persisted in all cats. The large frequency of grooming induced by BC was significantly reduced. SKF 525A pretreatment was correlated with a significant increase in staring and quiet sitting and a failure of BC to increase activities such as rubbing, treading and kneading. But many other BC-induced behaviors showed no changes. The data demonstrated that particular BC-induced changes in cats are antagonized by SKF 525A. The behavioral suppression caused by SKF 525A is compatible with the involvement of active hepatic metabolites from BC.(ABSTRACT TRUNCATED AT 250 WORDS)

The presence of phenylethanolamine-N-methyl transferase in those brain-stem nuclei where methyldopa is thought to act suggested to us that methylepinephrine could be an active metabolite of methyldopa. We assessed the potential of methylepinephrine to be an active metabolite by determining its capacity to compete for ligands which bind to alpha-receptors in rat brain. Thus, the abilities of epinephrine, norepinephrine, methyldopa, (+/-)-methylnorepinephrine and (+/-)-methylepinephrine to compete for alpha 1-([3H]WB-4101) and alpha 2-([3H]clonidine) binding sites in rat brain wee compared. Order of potency at each binding site was: [3H]WB-4101 -- (-)epinephrine = (-)-norepinephrine greater than (+/-)-methylnorepinephrine greater than (+/-)-methylepinephrine much greater than (-)-methyldopa; [3H]clonidine -- (-)-epinephrine greater than (+/-)-methylnorepinephrine greater than (-)-norepinephrine greater than (+/-)-methylepinephrine much greater than (-)-methyldopa. Since it can compete with a specific ligand for alpha 2-receptors, methylepinephrine could be a centrally active metabolite of methyldopa.

Intracerebroventricular (i.c.v.) injection of histidyl-proline diketopiperazine [cyclo (His-Pro)], an active metabolite of thyrotropin-releasing hormone (TRH) in mice produced an antinociceptive effect in a dose-dependent manner as measured in four antinociceptive tests; tail-pressure, tail-flick, hot-plate and acetic acid writhing. The antinociceptive effect of cyclo (His-Pro) reached a maximum at 5 min and lasted for 30 min. The ED50 values for the tests were 760.0 (598.4-965.2), 540.0 (442.6-658.8), 595.0 (487.7-725.9) and 370.0 (286.8-477.3) nmol/mouse and the slope functions were 1.61, 1.56, 1.57 and 1.66, respectively. Pretreatment with naloxone (0.5, 2 and 8 mg/kg), an opioid antagonist, administered subcutaneously antagonized the antinociceptive effect of cyclo (His-Pro). TRH injected i.c.v. to mice also exerted a dose-dependent antinociceptive action as measured in three antinociceptive tests; tail-pressure, hot-plate and acetic acid writhing. The antinociceptive effect of TRH was only seen at 5 min. The ED50 values for each test were 112.0 (47.5-264.3), 19.2 (10.4-35.5) and 0.2 (0.1-0.4) nmol/mouse and the slope functions were 8.89, 4.14 and 3.94, respectively. TRH was without effect in the tail-flick test. In contrast to cyclo (His-Pro), TRH-induced antinociception was not antagonized by pretreatment with naloxone (0.5, 2 and 8 mg/kg). The data suggest that the two peptides have a different mechanism of antinociceptive action in relation to the involvement of the opioid system in the central nervous system.

Pharmacological effects of Δ 8 -tetrahydrocannabinol ( Δ 8 -THC) and its metabolites, 11-hydroxy- Δ 8 -THC, 11-oxo- Δ 8 -THC and Δ 8 -THC-11-oic acid were compared using mice. The cataleptogenic effect of the 11-hydroxy and 11-oxo metabolites was 5 and 1.5 times greater respectively, than that of the parent compound. The hypothermic effect of Δ 8 -THC, 11-hydroxy- Δ 8 -THC and 11- oxo - Δ 8 -THC was almost equivalent in both potency and duration at a dose of 10 mg/kg i.v., but the metabolites exhibited a somewhat higher potency and longer duration that the parent compound at a dose of 5 mg/kg i.v. In addition, 11-hydroxy- and 11-oxo- Δ 8 -THC were more active to prolong pentobarbital-induced sleeping time than Δ 8 -THC. In spite of the loss of cataleptogenic and hypothermic effects, Δ 8 THC-11-oic acid slightly prolonged pentobarbital-induced sleeping time at a dose of 10 mg/kg i.v. The LD 50 s (i.v.) with their 95% confidence limits of Δ 8 -THC, 11-hydroxy- Δ 8 -THC and 11-oxo- Δ 8 -THC were estimated to be 27.5 (23.1–32.7), 110.0 (79.1–152.9) and 63.0 (54.5–72.8) mg/kg (P Δ 8 -THC-11-oic acid. These results indicate that both 11-hydroxy- and 11-oxo- Δ 8 -THC can be categorized as active metabolites of Δ 8 -THC. Further studies are necessary, however, to clarify whether or not these metabolites contribute, at least in part, to the effect of Δ 8 -THC on biological systems in vivo.

The influence of thyrotropin releasing hormone (TRH) and its postulated metabolite histidyl-proline diketopiperazine (HPD) on sedation and analgesia produced by delta 9-tetrahydrocannabinol (THC) was investigated in mice. Intracerebral administration of HPD (10 microgram) by itself decreased motor activity, whereas, TRH (10 microgram) was without any effect. THC (10 and 20 mg/kg) given i.p. decreased the motor activity and this effect was antagonised by prior treatment with TRH or HPD. Administration of THC produced analgesia as evidenced by delay in the tail-flick reaction time to a thermal stimulus. Pretreatment with TRH or HPD also antagonized the THC-induced analgesia. THC-induced analgesia was not modified by s.c. pretreatment with naloxone, an opiate antagonist. TRH or HPD pretreatment had no effect on the distribution of total radioactivity representing THC and its metabolites in brain and plasma. These studies indicate that TRH and HPD can antagonize THC-induced sedation and analgesia and that HPD may be an active metabolite of TRH. These effects were observed without altering the distribution of the drug and metabolite in brain and plasma, and therefore their effect may involve their direct actions on the central nervous system. Finally THC induced analgesia does not seem to involve opiate receptors and their involvement in the mechanism by which TRH or HPD antagonize THC-induced analgesia is unlikely.