Larry R. Steranka

Bradykinin receptor antagonists

Bradykinin and its active metabolites are produced at the sites of their actions by kallikreins. They potently elicit a variety of biological effects: hypotension, bronchoconstriction, gut and uterine contraction, epithelial secretion in airway, gut, and exocrine glands, vascular permeability, pain, connective tissue proliferation, and eicosanoid formation. These effects are mediated by at least two broad classes of receptors. The most common is the B2 subtype. The Stewart and Vavrek peptides characterized by a DPhe7 substitution have provided powerful tools for study of bradykinins actions by competitively and specifically blocking bradykinin B2 receptors. The significance of kinins in certain human diseases is being explored using these new tools and potential therapeutic agents. At present, human clinical trials are underway to test the usefulness of bradykinin receptor antagonists in the symptoms of the common cold and in the pain associated with severe burns. Trials for use in asthma will be initiated in 1990.

Immediate and long-term effects of p-chloroamphetamine on brain amines.

The administration of p-chloroamphetamine (p-CA) to rats induces a complex pattern of changes in brain amines and behavior. The immediate effects are relatively nonspecific, involving changes in serotonergic, noradrenergic, and dopaminergic neuronal systems that mediate a variety of behavioral effects. However, the persistent, long-term effects are specific to serotonin [Shydroxytryptamine (5-HT)I neurons. Moreover, the mechanism that mediates the long-term reduction of brain 5-HT is different from that responsible for the immediate decrease in this amine. Therefore, these two phases of p-CAs action will be considered separately.

Long-term effects of continuous exposure to amphetamine on brain dopamine concentration and synaptosomal uptake in mice

Continuous release of p-chloroamphetamine from subcutaneously implanted minipumps for 3 days decreased brain 5-hydroxytryptamine, but not norepinephrine or dopamine, when analyzed 2 weeks later. In contrast, 2 weeks after infusion of amphetamine, whole brain dopamine but not 5-hydroxytryptamine or norepinephrine, was decreased. Also, the high affinity uptake of [3H] dopamine was markedly and selectively reduced in striatal synaptosomes. Thus, neurotoxicity is apparently not unique to halogenated amphetamine derivatives, but is produced by amphetamine itself if administered continuously.

Antagonists of B2 bradykinin receptors.

Bradykinin and its active metabolites, produced by kallikreins at their sites of action, potently elicit a variety of biological effects: hypotension, bronchoconstriction, gut and uterine contraction, epithelial secretion in airway, gut, and exocrine glands, vascular permeability, pain, connective tissue proliferation, cytokine release, and eicosanoid formation. These effects are mediated by at least two broad classes of receptors. The most common is the B2 subtype. The availability of competitive antagonists of B2 receptors has provided powerful tools for the study of bradykinins actions. The significance of kinins in certain human diseases is being explored by using these agents as potential therapeutic agents. Human clinical trials are under way to test the usefulness of bradykinin receptor antagonists to treat symptoms of the common cold and the pain associated with severe burns. Trials are also being comtemplated for use in treatment of asthma.—Steranka, L. R.; Farmer, S. G.; Burch, R. M. Antagonists of B2 bradykinin receptors. FASEB J. 3: 2019‐2025; 1989.

Long-term effects of fenfluramine on central serotonergic mechanisms ☆

Abstract Consistent with results of previous studies, the present data indicate long-term changes in serotonergic neurons following a single injection of fenfluramine in rats. The brain levels of 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), tryptophan hydroxylase (TPH) activity and the synaptosomal uptake of 3H-5-HT were markedly reduced 2 weeks after 40 mg/kg of fenfluramine, and, with the exception of tryptophan hydroxylase activity, remained significantly decreased for at least 2 months. Although substantial recovery was evident between 2 weeks and 2 months, regional studies revealed marked reductions in some areas at 2 months. The largest reductions in 5-HT, 5-HIAA and 5-HT uptake were found in the hippocampus and cortex. The pattern of reduction of TPH was different, with the largest reductions in midbrain and limbic forebrain. The administration of 10 mg/kg of fluoxetine, a specific inhibitor of 5-HT uptake, prior to fenfluramine, prevented the long-term decreases in 5-HT, 5-HIAA and 5-HT uptake capacity, but not the decrease in TPH. These data suggest that the reduction in enzyme activity is mediated by a mechanism different from that responsible for the decreases in the other parameters. Thus, it is likely that the long-term effects of fenfluramine on brain 5-HT neurons can not be explained by a single mechanism such as an irreversible cytotoxic action.

Role of Neuronal Signal Input in the Down-Regulation of Central Noradrenergic Receptor Function by Antidepressant Drugs

Abstract: Rats with unilateral lesions of the locus coeruleus were used to study the role of norepinephrine (NE) signal input in the down‐regulation by antidepressants of the noradrenergic cyclic AMP‐generating system in the cortex. Chronic administration of both desipramine (blockade of NE reuptake) and iprindole (no blockade of NE reuptake) reduced the cyclic AMP response to NE on the nonlesioned side, but had little or no effect on the lesioned side. The results indicate that NE signal input and thus the formation of the NE‐receptor complex are prerequisites for inducing noradrenergic subsensitivity.

Species differences in the rate of disappearance of fenfluramine and its effects on brain serotonin neurons

Abstract A single injection of fenfluramine, the m -trifluoromethyl- N -ethyl derivative of amphetamine, produces long-term decreases in brain levels of 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), tryptophan hydroxylase activity (TPH), and the synaptosomal uptake of [ 3 H]-5-HT in rats. In order to test the extent to which similar effects occur in mice, rats and mice were injected with various doses of fenfluramine and killed 2 weeks later for the determination of 5-HT and 5-HIAA in one half of the brain, 5-HT uptake in the hippocampus from the other half, and TPH in the remaining tissue. A dose of 20 mg/kg produced marked reductions in 5-HT and 5-HIAA in rats, while in mice significant reductions in both of these parameters were observed only after 80 mg/kg. TPH was diminished after 60 mg/kg in rats but not after any dose up to 80 mg/kg in mice. A 58 percent reduction in 5-HT uptake occurred after 40 mg/kg in rats and 80 mg/kg in mice. Complete recovery was evident by 2 months after the administration of fenfluramine to mice when, as indicated by previous reports, marked effects are still present in rats. These results demonstrate that mice are substantially less sensitive than rats to the long-term effects of fenfluramine on central 5-HT systems. In addition, the half-lives of fenfluramine and its active metabolite, norfenfluramine. were much shorter in mice than in rats. Thus, the rates of disappearance of fenfluramine and norfenfluramine are correlated with, and perhaps determine, the extent to which the administration of fenfluramine produces longterm, neurotoxic effects on central 5-HT mechanisms in rats and mice.

A pivotal role for serotonin (5HT) in the regulation of beta adrenoceptors by antidepressants: reversibility of the action of parachlorophenylalanine by 5-hydroxytroptophan

An acute reduction in the synaptic availability of serotonin (5HT) by p-chlorophenlalanine (PCPA) nullifies the decrease in the density of cortical beta adrenoceptors caused by desipramine (DMI) but does not appreciably alter the attenuation of the norepinephrine (NE) sensitive adenylate cyclase. The analysis of competition-binding curves of [3H]-dihydroalprenolol shows that the affinity of the agonist (−)-isoproterenol for cortical beta adrenoceptors is profoundly reduced following PCPA. This reduction in agonist affinity is enhanced by DMI. Resupplying 5HT by by-passing trptophan hydroxylase inhibition, by administering 5-hydroxytryptophan, converts a DMI non-responsive to a DMI responsive beta adrenoceptor population and shifts the markedly decreased agonist affinity towards the affinity values found in control preparations. The results demonstrate the pivotal role of 5HT in the regulation of the density and agonist affinity characteristics of cortical beta adrenoceptors and contribute to the scientific basis of the ‘serotonin-norepinephrine link hypothesis’ of affective disorders.

Effect of cysteine on the persistent depletion of brain monoamines by amphetamine, p-chloroamphetamine and MPTP.

The administration of L-cysteine (500 mg/kg i.p.) 30 min before and 5 h after the administration of (+)-amphetamine sulfate markedly attenuated the persistent decreases in striatal dopamine (DA), dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in rats one week after the administration of a single dose of amphetamine (9.2 mg/kg i.p.) to iprindole-treated animals and in mice one week after the last of four daily injections of amphetamine (30 mg/kg i.p.). Cysteine prevented the persistent decreases in striatal serotonin (5HT) and 5-hydroxyindoleacetic acid (5HIAA) one week after the administration of p-chloroamphetamine to rats, but failed to alter the persistent decreases in striatal DA, DOPAC and HVA in mice one week after the last of four daily doses of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 30 mg/kg s.c.). The results suggest that the mechanisms by which amphetamine and p-chloroamphetamine, but not MPTP, produce persistent depletions of striatal monoamines involve the generation of neurotoxic electrophilic intermediates which can be inactivated by the administration of cysteine.

D-Phe7-substituted peptide bradykinin antagonists are not substrates for kininase II

The bradykinin receptor antagonists [D-Phe7]bradykinin, D-Arg[Hyp3,D-Phe7]bradykinin and D-Arg[Hyp3,Thi5,8,D-Phe7]bradykinin were tested for their ability to serve as substrates for kininase II (angiotensin converting enzyme) purified from rabbit lung. By HPLC, the peptides were not measurably degraded over 30 minutes. Under identical conditions, bradykinin was completely degraded to bradykinin (1-7). When hippuryl-His-Leu was used as a substrate for kininase II, the D-Phe7-substituted bradykinins acted as weak noncompetitive inhibitors. While the peptides were poor substrates for kininase II, they were short-lived when injected intravenously. D-Arg[Hyp3,D-Phe7]bradykinin was completely degraded to small fragments in less than 2 minutes. In diluted serum in vitro, a single product was observed with elution consistent with loss of arginine, suggestive of metabolism by kininase I.