Biomedical subjects
S Spector
Publications and source records attributed to S Spector.
Differentiation of Friend erythroleukemia cells induced by benzodiazepines.
Many benzodiazepines of the type whose receptors are found in peripheral tissues cause the differentiation of cultured Friend erythroleukemia cells. The maximal level of induction of hemoglobin synthesis is 10-80% of the cells, depending on the compound tested. The induction is concentration and time dependent, requiring micromolar amounts of the drugs and about 5 days of treatment for full expression. The time course is very similar to that observed for a well-studied inducer, dimethyl sulfoxide. The affinities of the agents for the peripheral-type benzodiazepine binding site are not correlated with their capacity to induce differentiation. Also, the biological effect is stereospecific since the (3S) stereoisomer Ro11 -6896 is at least an order of magnitude more potent than its (3R) enantiomer, Ro11 -6893. The benzodiazepine effect exhibits definite structure-activity relationships. A 1-methyl group is an absolute requirement, although this is not sufficient in itself. Hydroxyl and methoxyl groups at the 4' position enhance the biological activity, but 4'-chloro decreases it. Substitutions at the 2', 6', and 4 positions also decrease the biological activity, as does the lack of a 2-carbonyl group. These data suggest that the benzodiazepines act in a specific manner to induce the differentiation of Friend erythroleukemia cells.
Benzodiazepines that bind at peripheral sites inhibit cell proliferation.
[3H]Ro5-4864 binds to mouse thymoma cells in a specific, saturable, and reversible manner. Scatchard analysis shows a single class of binding sites with a Kd of 4.4 nM and a Bmax of 477 fmol per 10(6) cells. This benzodiazepine binding site is of the peripheral type, based on the relative potencies of Ro5-4864 and clonazepam in competing for [3H]diazepam binding. Fifteen benzodiazepines that bind to this site with affinities ranging from 6 nM to 1 microM also reversibly inhibit the proliferation of thymoma cells in culture at the micromolar dose range. There is a strong positive correlation (r = 0.85) between the binding constants of these compounds for the peripheral-type sites and their ED50 in inhibiting the uptake of [3H]thymidine into the cells. These sites may be involved in the regulation of thymoma cell proliferation.
Interaction of calcium channel blockers with non-neuronal benzodiazepine binding sites.
The ability of calcium channel blockers to displace the binding of benzodiazepine ligands was investigated in rat heart, kidney, and brain. The dihydropyridine calcium channel blockers nifedipine and nitrendipine displaced the binding of the non-neuronal-site ligand [3H]Ro5-4864, but not that of the neuronal-site ligands [3H]flurazepam or [3H]clonazepam. The inhibition was competitive, with Ki values in the micromolar range. Other calcium channel blockers--i.e., verapamil and diltiazem--were inactive at both sites. Thus, non-neuronal benzodiazepines bind to a class of sites that also binds dihydropyridines. This implies a role for benzodiazepines in the mediation of calcium-dependent phenomena.
Melatonin in serum and the pineal of spontaneously hypertensive rats.
Involvement of melatonin in the blood pressure regulation as an endogenous central hypotensive factor has been suggested in rats and in man. We studied the relationship between melatonin and the development of hypertension in 5- and 15-week-old spontaneously hypertensive (SHR) and normotensive Wistar Kyoto (WKY) rats, by measuring serum and the pineal concentrations with a sensitive and specific radioimmunoassay coupled with a novel extraction method. Serum melatonin concentration at midnight in young SHR rats was significantly higher than that in age-matched WKY rats (P less than 0.01), whereas it was decreased in the adult SHR (P less than 0.01). No such differences were observed at noon. Pineal content of melatonin at midnight in 5-week-old SHR rats was lower than in age-matched WKY rats (P less than 0.01). These data demonstrate that melatonin in the nocturnal serum of SHR rats is elevated at prehypertensive stage while it is decreased after the development of hypertension. The role of melatonin in the hypertensive process in SHR rats requires further study.
Structural requirements for the binding of benzodiazepines to their peripheral-type sites.
The DS19 Friend erythroleukemia cells possess peripheral-type benzodiazepine recognition sites that are similar to those characterized in other tissues and cells. These sites recognize only specific and well-defined ligand structures. In order for the benzodiazepines to bind to these sites, an alkyl group of 3 carbons or less is required at position 1, together with a carbonyl group at position 2. In addition, the binding affinity is enhanced by 4'-halogen or 4'-methoxy substituents, and by 7- or 2'-halogens. However, the affinity is decreased by substitutions at position 3 or 4. Central-type benzodiazepine binding in brain, on the other hand, is not affected by 1-alkyl groups and is inhibited by 4'-substitutions. Furthermore, unlike the peripheral-type site, the brain receptor is stereo-selective. These results demonstrate that the two binding sites are fundamentally different in their recognition of benzodiazepine ligands.
Structure-activity requirements for hypotension and alpha-adrenergic receptor blockade by analogues of atropine.
The hypotensive action of various antimuscarinic compounds structurally related to atropine was studied in conscious, unanesthetized rats. The alpha-adrenolytic activity of these agents was assessed both in vivo (blockade of norepinephrine-induced pressor response) and in vitro (displacement of [3H]WB-4101 binding). Benztropine, homatropine and hyoscyamine caused hypotension and produced alpha-adrenergic receptor blockade similar to atropine. Other analogues were either inactive (atroscine, scopolamine, tropic acid and tropine) or evoked nonspecific changes in blood pressure and lacked alpha-adrenolytic activity (benactyzine, eucatropine, methylatropine, methylhomatropine and methylscopolamine). Based on these data, we propose the following structure-activity relationship for hypotension and alpha-adrenolytic activity: (a) the tropine moiety is inactive unless it is attached to another group by an ester linkage, (b) chemical modification of the tropine moiety, including quaternization, decreases potency, (c) the d-stereoisomer appears to be more potent than the corresponding 1-form.
The binding of [3H]-diazepam to guinea-pig ileal longitudinal muscle and the in vitro inhibition of contraction by benzodiazepines.
1--The longitudinal muscle-myenteric plexus strip preparation of the guinea-pig ileum was used to study the binding of [3H]-diazepam and the effect of benzodiazepines on its contraction. 2--Scatchard analysis of binding indicated a single class of binding sites with KD = 43 nM and Bmax = 229 fmol/mg prótein. Binding was of peripheral type based on the much greater binding affinity of Ro5-4864 as compared to clonazepam. Binding of [3H]-diazepam reached equilibrium at 10 min and dissociated rapidly (T1/2 = 1.3 min). The KD derived from the rate constants agreed with that from the Scatchard analysis. 3--Benzodiazepines produced a dose-dependent decrease in the electrically induced contractions of the longitudinal muscle strip, but their potencies in this effect did not correlate with their binding affinities. 4--Diazepam antagonized the contractions of the longitudinal muscle strip induced by K+, Ca2+, histamine and carbachol. The inhibition of Ca2+-induced contractions was reversed by increasing the concentration of Ca2+ in the medium.
[3H]Diazepam binding sites on rat heart and kidney.
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Placental theophylline transfer in pregnant asthmatics.
The newborn's potential for xanthine toxic reactions from placental transfer of theophylline or caffeine conversion from theophylline was studied in 12 newborns of asthmatic mothers. Our results showed that maternal and cord theophylline levels were not notably different, yet heelstick levels tended to be higher than maternal levels. Newborn heelstick theophylline levels ranged from 2.3 to 19.6 microgram/mL with a mean of 10.5 microgram/mL. No caffeine conversion from theophylline was seen. Heart rate, Apgar scoring, and other parameters were not statistically different in babies whose mothers received theophylline compared with mildly asthmatic mothers not receiving theophylline and nonasthmatic mothers. Side effects in three babies with levels of theophylline greater than 10 microgram/mL were tachycardia and transient jitteriness. Newborns seem to tolerate theophylline levels corresponding to maternal levels without serious adversity; however, transient toxicity might occur with levels in the so-called therapeutic range.
Opioid polypeptides in guinea pig pancreas.
Extracts from guinea pig pancreas were found to contain high molecular weight enkephalin-containing polypeptides as well as free [Met]enkephalin and [Leu]enkephalin and the hexa-, hepta-, and octapeptides derived from proenkephalin. Within the limits of sensitivity of our assay (10 fmol/g of tissue), beta-endorphin and beta-lipotropin were undetectable.
Disposition of drugs in man by radioimmunoassay.
The disposition of morphine and clonidine was investigated by means of radioimmunoassay. After the administration of a single i.v. dose of morphine (10 mg/70 kg) to 10 men who had not received other drugs for two weeks preceding the study, a multiphasic decline in serum concentrations of morphine occurred. Detectable blood concentrations of morphine, or a metabolite, or of both, persisted for up to 96 hours after a single i.v. dose. The radioimmunoassay for clonidine is capable of detecting as little as 10 pg of clonidine. The antibody fails to bind 2,6-dichlorophenylguanidine, a known metabolite of clonidine, while the other metabolite, 4-hydroxyclonidine, was as potent as clonidine in displacing labeled clonidine from the antibody. The disposition of clonidine ion plasma and tissues could be monitored by radioimmunoassay.
Changes in platelet and renal benzodiazepine binding in spontaneously hypertensive rats.
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Central neurotransmitter receptors in hypertensive rats.
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Atropine lowers blood pressure in normotensive rats through blockade of alpha-adrenergic receptors.
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Studies on the hypotensive response to atropine in hypertensive rats.
The hypotensive effect of atropine sulfate was evaluated in conscious, unanesthetized hypertensive rats (spontaneously hypertensive rats and Sprague-Dawley rats made hypertensive by s.c. implantation of deoxycorticosterone acetate, tablets) and their respective normotensive controls. In all animals, atropine (5-50 mg/kg i.v.) caused an immediate and dose-dependent decrease in blood pressure. However, the hypotensive response was of greater magnitude and of longer duration in the hypertensive rats. Pretreatment of the animals with phentolamine prevented the hypotension by atropine. Plasma concentrations of atropine were determined by radioimmunoassay and were identical in both hypertensive and normotensive rats. Atropine blocked the pressor responses to norepinephrine and shifted the dose-response curve for norepinephrine to the right to a similar extent in spontaneously hypertensive and Wistar-Kyoto rats. Thus, atropine acts as a competitive antagonist of norepinephrine and this action underlies its hypotensive effect. The greater responsiveness of hypertensive rats is not due to a difference in pharmacokinetic handling of atropine nor to a greater degree of blockade of the norepinephrine pressor response in vivo, but may result from the altered sympathetic tone in the hypertensive rats. Atropine may, in addition to its action on alpha adrenergic receptors, affect other systems such as calcium handling. It is speculated that an effect on this system may contribute to the longer duration of response seen in hypertensive rats.
Tolerance to beta-adrenergics and results of rising dose studies of albuterol, isoproterenol and placebo.
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