Antinociceptive profile of sulfated CCK-8. Comparison with CCK-4, unsulfated CCK-8 and other neuropeptides.

The antinociceptive activity of sulfated cholecystokinin octapeptide (CCK-8-S) was characterized by comparison with two other endogenous forms, unsulfated CCK-8 (CCK-8-U) and the carboxyl tetrapeptide fragment (CCK-4) and two other peptides present in the gut and brain: bombesin and neurotensin. By the intracerebroventricular (i.c.v.) route, CCK-8-S was antinociceptive in the hot plate and phenylquinone-induced writhing assays, but CCK-8-U and CCK-4 were active only in the latter test. By systemic administration, CCK-8-S retained anti-writhing activity but CCK-8-U and CCK-4 did not. Therefore, CCK receptors in brain may be involved in the apparent antinociception produced by CCK-8-U and CCK-4. Bombesin produced potent antinociceptive activity, along with a distinct, head-scratching syndrome, in both the writhing and hot plate tests. Naloxone reversed bombesin-induced elevation of latencies of mouse jump but not the head-scratching syndrome, indicating that the analgesic effect in the hot plate test was independent of the scratching behaviour. Neurotensin, unlike CCK-8-S, elevated tail-flick latencies, and was more potent in the writhing than in the hot plate test. Several differences between CCK-8-S and opioid substances included the need for relatively large doses of naloxone to block the effects of CCK-8-S in the phenylquinone-induced writhing test and the lack of effect of CCK-8-S in the tail-flick test. Global sedation can account for some, but not all, of the effects of CCK-8-S.(ABSTRACT TRUNCATED AT 250 WORDS)

The neuropharmacology of various diazepam antagonists.

Recently, compounds which bind avidly to benzodiazepine binding sites have been shown to possess diazepam antagonist properties. For example, the benzodiazepine RO 15-1788 and the pyrazoloquinoline CGS 8216 can antagonize the anxiolytic, sedative, muscle relaxant and anticonvulsant properties of diazepam. The beta-carbolines have also been shown to antagonize several actions of diazepam. Other compounds including physostigmine, naloxone, bicuculline, picrotoxin, caffeine and theophylline, lack appreciable affinity for benzodiazepine binding sites but do antagonize at least some of the behavioral actions of diazepam. Their antagonist properties are probably the result of opposing pharmacological actions rather than direct receptor antagonism. Clinically, a potent safe diazepam antagonist could be used to reverse effects of diazepam overdose and to speed recovery of diazepam-treated patients after various out-patient procedures.

Synthesis and benzodiazepine binding activity of a series of novel [1,2,4]triazolo[1,5-c]quinazolin-5(6H)-ones.

Investigation of tricyclic heterocycles related to the 2-arylpyrazolo[4,3-c]quinolin-3(5H)-ones, structures with high affinity for the benzodiazepine (BZ) receptor, led to the synthesis of 2-phenyl-[1,2,4]triazolo[1,5-c]quinazolin-5(6H)-one, a compound with 4 nM binding affinity to the BZ receptor. Analogues were prepared to assess the importance of the 2-substituent and ring substitution in modifying activity. Several novel synthetic routes were designed to prepare the target compounds, including a two-step synthesis beginning with an anthranilonitrile and a hydrazide. Of the 34 compounds screened in this series, three compounds were found to be potent BZ antagonists in rat models. The leading compound, 9-chloro-2-(2-fluorophenyl) [1,2,4]triazolo[1,5- c]quinazolin-5(6H)-one (CGS 16228), showed activity comparable to that of CGS 8216 from the pyrazolo[4,3-c]quinoline series.

Antagonism of morphine analgesia by CCK-8-S does not extend to all assays nor all opiate analgesics.

The conditions under which CCK-8-S may block opiate-induced analgesia were examined in detail. A U-shaped dose-response relationship was observed for the ability of CCK-8-S to attenuate (by approximately 50%, at most) morphine-induced tail flick analgesia. The analgesic effects of morphine in the hot plate or acetic acid-induced stretching tests were not altered by CCK-8-S at doses that antagonized morphine in the tail flick test. Tail flick latency elevations induced by meptazinol, a putative mu-1 receptor agonist, were also attenuated by CCK-8-S according to a U-shaped dose-response relationship, but those induced by U-50,488, a kappa agonist, were not antagonized by CCK-8-S doses that attenuated morphine analgesia. Thus, the ability of CCK-8-S to antagonize opiate analgesia does not follow a conventional dose-response relationship, does not extend to all tests of analgesia and may not extend to all opioid drugs. Analgesia mediated by the mu-1 opioid receptor subtype may be more amenable to antagonism by CCK-8-S than that mediated by the kappa receptor subtype.

Unsulfated CCK-8 not blocked by proglumide or CR 1409 in the mouse abdominal irritant-induced stretching assay: possible central site of action.

Previous studies have shown that unsulfated cholecystokinin octapeptide (CCK-8-U) shares with the sulfated octapeptide (CCK-8-S) and the carboxyl-terminal tetrapeptide (CCK-4) the ability to block abdominal irritant-induced stretching when administered intracerebroventricularly. The effects of CCK-8-U, however, are not naloxone-reversible and do not appear upon systemic administration. To assess the hypothesis that the antistretching effects of CCK-8-U are mediated by central-type (CCK-B), rather than peripheral-type (CCK-A) receptors, the present experiments examined the reversal of these effects by CR 1409, a CCK receptor antagonist with in vitro selectivity for CCK-A receptors, and by proglumide. Both antagonists, when administered ICV, blocked the antistretching effects of CCK-8-S and CCK-4 (ICV), but not those of CCK-8-U. CR 1409 was approximately 40 times more potent against CCK-8-S by the ICV route than subcutaneously, indicating a likely action on CCK-A receptors in the brain. The effects of morphine, bombesin and neurotensin (ICV) were not blocked by CR 1409, indicating specificity for CCK receptor-mediated effects. The antistretching effects of CCK-8-U do not appear to be mediated by CCK-A receptors, and the possibility of a CCK-B receptor site of action must be considered.

Angiotensin-converting enzyme inhibitors potentiate the analgesic activity of [Met]-enkephalin-Arg6-Phe7 by inhibiting its degradation in mouse brain.

The proteolytic degradation of the enkephalin-containing heptapeptide Tyr-Gly-Gly-Phe-Met-Arg-Phe (YGGFMRF) was investigated by incubating the peptide with synaptic membranes from mouse whole brain and characterizing the formed products. The degradation products were derivatized with 4-dimethylaminoazobenzene-4'-isothiocyanate and then analyzed by high pressure liquid chromatography and by amino-terminal analysis. The incubation of YGGFMRF with synaptic membranes yielded YGGFM and RF as the degradation products. The angiotensin-converting enzyme (ACE) inhibitors, MK-422 and captopril, potently inhibited the formation of YGGFM and RF with IC50 values of 8 nM and 95 nM, respectively. The "enkephalinase A" inhibitor, thiorphan, weakly inhibited this dipeptidyl carboxypeptidase activity with an IC50 greater than 1 microM. YGGFMRF, MK-422, captopril, and thiorphan all produced a dose-dependent analgesic response in the mouse hot plate test when administered intracerebroventricularly. However, when subanalgesic doses of inhibitors were co-administered with a subanalgesic dose of YGGFMRF, only the ACE inhibitors, MK-422 and captopril, potentiated the analgesic response of the peptide. These data provide in vitro and in vivo evidence that ACE is the primary enzyme involved in the proteolytic degradation of YGGFMRF in the mouse brain.

Differential antagonism by proglumide of various CCK-mediated effects in mice.

Proglumide has been reported to antagonize sulfated cholecystokinin octapeptide (CCK-8) in peripheral tissue and in neurophysiological single unit preparations. The present studies attempt to extend this reported antagonism to several actions of CCK-8 in mice in vivo. For comparison with proglumide, the antagonist activity of naloxone against CCK-8 has also been evaluated. Proglumide (150 mg/kg) antagonizes hot plate latency elevations produced by a moderate (0.17 mg/kg s.c.) dose of CCK-8, but not that by a higher (1.0 mg/kg) dose. The effects of intracerebroventricularly (i.c.v.) administered CCK-8 (0.3 micrograms) are also blocked by proglumide i.p. or i.c.v. Naloxone (0.5 mg/kg s.c.) significantly antagonizes the elevated hot plate latencies induced by CCK-8 i.c.v. (3.0 micrograms) and s.c. (3.0 mg/kg). Proglumide antagonizes the motor activity suppressant effects of CCK-8, but only at a high proglumide dose (150 mg/kg i.p.) and low CCK-8 doses. Naloxone (3.0 mg/kg) is not effective against CCK-8 in motor activity. The hypothermia induced by CCK-8 cannot be antagonized either by proglumide at doses up to 150 mg/kg i.p. or by naloxone at doses up to 3.0 mg/kg s.c. In vivo, proglumide may be considered as a weak antagonist of CCK-8 and the possibility exists that various actions of CCK-8 may be differentiated by their relative responsiveness to proglumide-induced antagonism.