Effects of endomorphin-2 on arterial blood pressure and pain threshold in spontaneously hypertensive rats and modification of these effects by beta-funaltrexamine and nor-binaltorphimine.

The effects of intracerebroventricular (icv) administration of endomorphin-2 (E2) on arterial blood pressure and pain threshold in spontaneously hypertensive rats (SHR) and modification of these effects by K [OP2] and mu [OP3] opioid receptors antagonists were investigated. Endomorphin-2 administrated icv in doses of 8, 16 and 32 mcg produced dose-dependent analgesic and hypotensive effect. In SHR decrease in blood pressure amounted 2.667, 4.0 and 6.534 kPa, respectively. Pain threshold increased by 1.7, 3.6 and 8.9 (g x 10). In Wistar Kyoto (WKY) strain, being the normotensive controls, E2 in doses of 8 and 16 mcg decrease in blood pressure was less pronounced and amounted 1.200 and 1.467 kPa, respectively, whereas the pain threshold increased by 7.2 and 10.4 (g x 10), respectively. Both E2 effects were antagonized by equimolar icv doses of beta-funaltrexamine (beta-FNA). Equimolar doses of nor-binaltorphimine (nor-BNI) attenuated analgesic action of E2, but were without hypotensive action produced by E2. A strong correlation between drop in blood pressure and increase in pain threshold observed in the SHR and WKY strains after icv administration of E2, indicate close interaction between systems responsible for pain perception and blood pressure control.

Paradoxical effects of intracerebroventricular low-dose opioid antagonists in SHR with chronic pain.

The aim of our study was to investigate the effect of intracerebroventricular (i.c.v.) administration of very low doses of opioid antagonists on the pain threshold, arterial blood pressure and body temperature of spontaneously hypertensive rats (SHR) with chronic pain. We found that low doses of i.c.v. administered naloxone hydrochloride (0.3 microg) or naloxone methiodide (0.4 microg) produce paradoxical hypoalgesia. Similar results were not observed following i.c.v. administration of nor-binaltorphimine (0.6 microg). A paradoxical increase in the severity of hypertension followed i.c.v. opioid antagonist administration. This suggests an involvement of the opioid system in the mechanisms of blood pressure control. The paradoxical results obtained both for pain threshold and blood pressure after low doses of some opioid antagonists seem to confirm the role played by opioid autoreceptors in these effects. Existence of autoreceptors is suggested. Results obtained following i.c.v. administration of nor-binaltorphimine also suggest a role for the kappa autoreceptor (OP2) in the regulatory mechanisms of thermoregulation.

Spinal co-administration of peptide substance P antagonist increases antinociceptive effect of the opioid peptide biphalin.

Intrathecal injection of 0.25 micrograms of undecapeptide substance P antagonist (SPA) produced transient antinociception with a peak effect at 5 min. Increasing the SPA dose resulted in neurotoxicity. Intrathecal injection of the opioid peptide biphalin (BIP) produced antinociception for over 3 hrs without neurotoxicity. Co-administration of SPA (at subtoxic doses) increased BIP's antinociceptive effect. Naltrexone reversed analgesia due to BIP alone as well as after BIP+SPA.

Synthesis of enkephalin analogs. Part VI. N,N-disubstituted derivatives.

Synthesis of four new N,N-disubstituted derivatives of enkephalin analogs: All2Tyr-DMet-Gly-Phe-epsilon Ahx-OMe 5, Bu2Tyr-DMet-Gly-Phe-epsilon Ahx-OMe 6, All2Tyr-DMet-Gly-Phe-epsilon Ahx-epsilon Ahx-OMe 11 and Bu2Tyr-DMet-Gly-Phe-epsilon Ahx-epsilon Ahx-OMe 12 is reported. they were tested for agonistic and antagonistic activity. Compound 5 is a little more potent agonist (IC50 = 1.9 x 10(-7) M/l, GPI) than compound 6(IC50 = 7.2 x 10(-7) M/l, GPI). They both are highly selective to mu receptor, because they show no trace of activity to delta receptor in concentration up to 10(-5) M/l. Compound 11 and 12 are less active and not selective as agonists. None of these compounds showed antagonistic activity.

Synthesis of enkephalin analogs. Part V. N-monosubstituted derivatives.

Synthesis of four new derivatives of enkephalin analogs: H-BuTyr-DMet-Gly- Phe-epsilon Ahx-OH 9, H-BzlTyr-DMet-Gly-Phe-epsilon Ahx-OH 12, H-Butyr-DMet-Gly- -Phe-epsilon Ahx-epsilon Ahx-OH 15 and H-BzlTyr-DMet-Gly-Phe-epsilon Ahx-OH is reported. They were examined for agonistic, antagonistic and analgesic activity. Compound 12 is the most potent among investigated peptides. Its agonistic activity in vitro is 7.85 x 10(-8) M/l (GPI) and 9.5 x 10(-7) M/l (MVD). None of the peptides showed antagonistic activity. Only compound 12 showed weak, not dose-dependent analgesic activity in rats.

The synthesis and pharmacological activity of two new derivatives of benzazocine.

Two new derivatives of benzazocine of anticipated analgesic action were synthesized. Pharmacological investigations were carried out to confirm their analgesic activity, affinity to the opiate receptor and potential antagonistic properties.

Synthesis and biological evaluation of human preproenkephalin (100-111) and its analogs.

The dodecapeptide sequence, Tyr-Gly-Gly-Phe-Met-Lys-Arg-Tyr-Gly-Gly-Phe-Met (BI), which is totally conserved in the primary structures of human, bovine, rat and toad preproenkephalins, has been synthesized by the solid-phase method. Coupling reactions were achieved by using symmetrical anhydrides of tert.-butyloxycarbonylamino acids performed with N-tert.-butyl,N'-methylcarbodiimide. 6-Arg and 7-Lys analogs have also been obtained. The peptides show opiate activity in both GPI and MVD assay, and possess antinociceptive properties as estimated by the hot-plate test in mice when applied intracisternally.

The effects of D-phenylalanine and its derivatives on enkephalin degradation in vitro: relation to analgesia and attenuation of the morphine withdrawal syndrome.

The effect of D-Phenylalanine (D-Phe), putative carboxypeptidase A inhibitor and its four derivatives (T1-T4) on analgesia, development of tolerance and physical dependence to morphine, and on degradation of both exogenous and endogenous enkephalins was investigated. Systemic administration of either D-Phe or its derivatives produced naloxone-reversible analgesia in the hot-plate test in mice. Naloxone-precipitated morphine withdrawal syndrome was attenuated in mice after systemic subacute administration (7 days, 1.2 mmol/kg, sc) of D-phe derivatives, the development of tolerance to morphine being unchanged. In the presence of either D-Phe or its derivatives in incubation mixture (up to 10(-3) mol/l) the hydrolysis of exogenous 3H-Met5-and 3H-Leu5-enkephalin in striatal homogenates was slightly inhibited. Moreover, the addition of D-Phe or its derivatives seemed to increase the per cent of recovered endogenous Met5-enkephalin released from veratridine-depolarized striatal particles. In contrast, bestatin, an amino-peptidase inhibitor, and a mixture of dipeptides (Tyr-Tyr, Leu-Leu, Leu-Gly) markedly inhibited degradation of both endogenous and exogenous enkephalins in vitro. The results obtained in this study suggest that that pharmacological activity of D-Phe is not directly related to the endogenous opiate system.

Modification of morphine withdrawal: effect of tuftsin, [Lys4]-tuftsinyltuftsin, tetrapeptide fragment (1-4) of substance P and its amide.

Withdrawal behavior in morphine-dependent rats precipitated by naloxone was attenuated after pretreatment with the tetrapeptide tuftsin and to some extent by its synthetic derivative [Lys4]-tuftsinyltuftsin. The tetrapeptide fragment (1-4) of Substance P was ineffective in suppressing morphine-withdrawal behavior, whereas its C-amide exerted only weak action. Possible involvement of an immunological mechanism is discussed.

The effects of apamin in rats with pretrigeminal or high spinal transsection of the central nervous system.

Rats were injected in one lateral cerebral ventricle (i.c.v.) with apamin (100 ng per animal). The resulting desynchronisation pattern in the electrocorticogram (ECoG) and the symptoms of poisoning were monitored before and after transsection at different levels, and following morphine. Apamin acts primarily on the brain stem and spinal cord, i.e. structures possessing a sensory input, and then indirectly on the higher integrating systems. There is no general parallelism between receptor density and locus of action.

Structure-activity studies of dermorphin. The role of side chains of amino acid residues on the biological activity of dermorphin.

Seventeen analogues of dermorphin were synthesized and bio-assayed to determine the influence of side chains of the individual amino acid residues forming the sequence of dermorphin on the biological activity of this opioid peptide. Syntheses were carried out using solid-phase procedure, and the analogues obtained were purified by gel filtration on Sephadex G-10. Biological activities determined in guinea pig ileum (GPI) and mouse vas deferens (MVD) tests showed that the N-terminal tetrapeptide is responsible for the activity of dermorphin. Substitutions in the C-terminal fragment, particularly in position 5, for other amino acid residues results in substantial differentiation towards mu and delta receptors.

Structure-activity studies of dermorphin. Synthesis and some pharmacological data of dermorphin and its 1-substituted analogues.

Dermorphin and its analogues substituted at position 1 by N-acetyltyrosine, O-methyltyrosine, phenylalanine, D-phenylalanine, or alanine were obtained by solid-phase peptide synthesis. Their pharmacological effects were studied in vitro by the guinea pig ileum method and in vivo by the hot plate method, and the results were compared with those of morphine. The most pronounced activity was shown for dermorphin. A radioreceptor study showed a moderate affinity of dermorphin and its Tyr(Me)1 analogue for the opiate receptor sites from striatal homogenates.

Effect of intracerebral administration of substance P on body temperature.

Effects of substance P (SP) administered into anterior hypothalamus or lateral ventricle on body temperature were investigated. When administered into lateral ventricle in doses from 20 to 2000 ng SP failed to influence body temperature. Application of SP into anterior hypothalamus in the same doses (20-2000 ng) resulted in a mild but insignificant increase in body temperature. No visible changes in the behavior of the rats, except slight sedation after the highest dose administered into lateral ventricle, were observed after application of SP. These results suggest that SP does not play any essential role in thermoregulation.

Lack of postulated opiate-receptor antagonistic properties of compound 48/80.

The previously suggested opiate-receptor antagonistic properties of compound 48/80 were checked both in vivo and in vitro. In electrically stimulated guinea pig ileum and rat vas deferens preparations compound 48/80 did not reverse the inhibition of the twitches caused either by morphine or Met5-enkephalin. In mice hot-plate test compound 48/80 did not decrease the analgesic activity of morphine. In radioreceptor studies compound 48/80 shows rather low affinity to the 3H-naloxone receptor sites from striatal homogenates of the rat, as compared to non-labelled naloxone and on the other hand it produced moderate, as compared to d-butaclamol, displacement of 3H-spiroperidol from rat striatal homogenates. The variation of composition of various polymers making compound 48/80 is suggested for explanation of obtained results.

Decreased myocardial and renal content of 3',5'-AMP in genetically hypertensive rats: effects of PGE2 and PGF2 alpha.

Levels of c-AMP in heart and kidney of genetically hypertensive rats (SHR) were significantly lower than in normotensive Wistar animals. PGE2, 10 microgram/kg ip markedly increased the concentration of c-AMP in heart and kidney of SHR animals, but was without effect on this cyclic nucleotide content in normotensive controls. PGF2, in doses up to 100 microgram/kg, did not influence the concentrations of c-AMP in heart and kidney of SHR animals. Alterations in adenylate cyclase--c-AMP system in genetic hypertension are discussed.

The effect of naloxone on cerebellar cGMP content. A possible GABA-antagonistic action?

Naloxone in high doses (60--240 mg/kg i.p.) produced a dose-dependent increase in cerebellar cGMP content of mice. The rise in cGMP content reached its maximum within 5 min and was of short duration. Short-lasting episodes of clonic seizures were noted after 240 mg/kg naloxone. Low doses of naloxone (5--10 mg/kg) had no effect on cerebellar cGMP content, but markedly potentiated the increase in cGMP induced by isoniazid. On the other hand, naloxone (5 mg/kg) partially antagonized the fall in cGMP elicited by diazepam, but had only a slight effect on the action of pentobarbital (30 mg/kg i.p.). These results support the assumption proposed by other authors that naloxone exerts GABA antagonistic effects aside from the potent opiate receptor antagonistic activity.

Baclofen and cerebellar cyclic GMP levels in mice.

Baclofen (3.5--10.0 mg/kg) dose dependently decreased cerebellar cyclic GMP content in mice correlated to a reduction in locomotor activity. Pretreatment with baclofen antagonized the rise in cerebellar cyclic GMP content induced by isoniazid and picrotoxin, but did not affect the increase in cyclic GMP following administration of pentetrazole. Baclofen had little effect on the arecoline-induced changes in cerebellar cyclic GMP content. Therefore, our data support the concept of gamma-aminobutyric acid agonistic properties of baclofen at presynaptic sites.