Itzchak Angel

[3H]GBR-12935 binding to the dopamine transporter is decreased in the caudate nucleus in Parkinson's disease

The specific binding of [3H]GBR‐12935 to membranes prepared from human caudate nucleus is saturable (Bmax 1.36 ± 0.18 pmol/mg protein), sodium dependent, and of high affinity (KD 2.34 ± 0.18 nM). Freezing of tissue from rat brain, or refrigeration followed by freezing, results in a small but significant (20%) decrease in specific [3H]GBR‐12935 binding when compared to the binding observed in fresh (nonfrozen) tissue, and this decrease may account, in part, for the differences in specific binding between rat and human brain membranes. Despite small differences in binding site density between fresh and frozen tissue there is a good correlation (r= 0.98; p < 0.01) between the potencies of a series of drugs in displacing specific [3H]GBR‐12935 binding to human caudate membranes and rat striatum as well as in inhibiting dopamine uptake in rat striatal synaptosomes (r= 0.96; p < 0.01). The specific binding of [3H]GBR‐12935 to membranes prepared from the caudate nuclei of patients with Parkinsons disease is decreased compared to membranes prepared from age‐and sex‐matched controls. These data suggest that [3H]GBR‐12935 binds in a sodium‐dependent fashion to the dopamine transport complex in human brain and that specific binding is decreased by a pathological degeneration of dopaminergic neurons to the caudate nucleus.

Inhibition of synaptosomal 5-[3H]hydroxytryptamine uptake by endogenous factor(s) in human blood

The inhibition of 5‐[3H] hydroxytryptamine uptake into rat forebrain synaptosomes by human plasma extracts was studied. Highly potent, small (M r < 10000), and heat stable factor(s) were found to inhibit 5‐[3H] hydroxytryptamine uptake specifically, reversibly and in a non‐competitive manner. The possible role of these factor(s) as endogenous modulators of serotonergic activity is discussed.

Glucose regulates [3H](+)-amphetamine binding and Na+K+ ATPase activity in the hypothalamus: A proposed mechanism for the glucostatic control of feeding and satiety

Binding sites for [3H](+)-amphetamine in the hypothalamus may mediate the anorectic actions of amphetamine and related phenylethylamines. To investigate further the role of these sites in the central control of appetite, the binding of [3H](+)-amphetamine to the hypothalamus and brainstem was measured following food deprivation and refeeding, the onset of genetic obesity, or the administration of 2-deoxy-D-glucose. Food deprivation for 24 to 72 hours reduced the Bmax for [3H](+)-amphetamine binding in the hypothalamus and brainstem but not in other brain areas or peripheral tissues. The decrease in hypothalamic and brainstem [3H](+)-amphetamine binding observed following food deprivation was time-dependent and rapidly reversed by brief refeeding with either rat chow or a 10% glucose solution. Moreover the changes in [3H](+)-amphetamine binding were highly correlated to corresponding alterations in blood glucose concentration. Furthermore, D-glucose, but not L-glucose increases the number of hypothalamic [3H](+)-amphetamine binding sites when administered in vivo or when added to hypothalamic slices in vitro. These data suggest that the [3H](+)-amphetamine binding site in the hypothalamus and (or) brainstem may be coupled to a central glucostat.

Characterization of [3H]Mazindol Binding in Rat Brain: Sodium‐Sensitive Binding Correlates with the Anorectic Potencies of Phenylethylamines

Abstract: Saturable low‐affinity binding sites for [3H]mazin‐dol have been demonstrated in crude synaptosomal membranes from rat brain using both a centrifugation and a fil‐tion assay. Studies on the regional distribution of these binding sites revealed that the hypothalamus and brainstem had the highest density of sites. Kinetic analysis of the binding of [3H]mazindol to hypothalamic membranes demonstrated a single class of noninteracting binding sites with an apparent affinity constant (KD) of 10.2 ± 0.7 μM and maximal number of binding sites (Bmax) of 786 ± 94 pmol/mg of protein. Specific [3H]mazindol binding was rapidly reversible, temperature sensitive, labile to pretreatment with proteolytic enzymes, and inhibited by physiological concentrations of sodium. In most peripheral tissues, such as the liver and kidney, very low levels of binding were observed; however, the adrenal gland had a relatively high density of sites. The potency of a series of anorectic drugs in inhibiting specific [3H]mazindol binding to hypothalamic membranes was highly correlated with their anorectic potencies in rats, but not with their motor stimulatory effects. These results suggest the presence of a specific drug recognition site in the hypothalamus that may mediate the anorectic activity of mazindol and related phenylethylamines.

In vitro stimulation of insulin release by SL 84.0418, a new alpha 2-adrenoceptor antagonist.

SL 84.0418 (2-(4,5-dihydro-1H-imidazol-2-yl)-1,2,4,5-tetrahydro-2-propyl-pyrrolo[3, 2,1- hi]-indole hydrocholoride) is a novel alpha 2-adrenoceptor antagonist which possesses anti-hyperglycaemic properties in vitro study, we tested its effects on insulin release from isolated mouse islets. In the presence of 15 mM glucose, SL 84.0418 produced a concentration-dependent (0.1-100 microM) increase of insulin release with a slightly higher potency than tolbutamide. SL 84.0418 antagonized the inhibition of glucose-induced insulin release caused either by the alpha 2-adrenoceptor agonist clonidine or by diazoxide, a selective opener of ATP-sensitive K+ channels in the beta-cell membrane. Its potency was greater on the inhibition by clonidine than on that by diazoxide, complete antagonism of the inhibition being achieved by 0.9 microM and 6 microM SL 84.0418 respectively. When alpha 2-adrenoceptors were blocked by the antagonist idazoxan, the low concentrations of SL 84.0418 (0.1-0.3 microM) no longer increased insulin release, whereas the effect of higher concentrations (> or = 1 microM) was not affected. SL 84.0418 (> or = 1 microM) inhibited 86Rb efflux from islets perifused with a medium containing 3 mM glucose, i.e. under conditions where many ATP-sensitive K+ channels are open. It also reduced the acceleration of 86Rb efflux that diazoxide caused in the presence of 6 mM glucose. Moreover, SL 84.0418 directly inhibited ATP-sensitive K+ currents measured in single beta-cells by the whole-cell mode of the patch-clamp technique.(ABSTRACT TRUNCATED AT 250 WORDS)

Site of action of anorectic drugs: Glucoprivic- versus food deprivation-induced feeding

Feeding induced by 2-deoxyglucose was compared with feeding induced by food deprivation in terms of antagonism by anorectic drugs and of anatomical site of action. Glucoprivic feeding was completely blocked by microinjection of amphetamine, fenfluramine, and mazindol into the paraventricular nucleus of the hypothalamus (PVN). Deprivation-induced feeding was not blocked by amphetamine, fenfluramine, or mazindol microinjected into the PVN. Neither the feeding induced by 2-deoxyglucose nor its reversal by amphetamine were blocked by pretreatment with the beta-adrenergic antagonist, propranolol. Amphetamine and fenfluramine blocked both glucoprivic- and deprivation-induced feeding when microinjected into the perifornical region of the lateral hypothalamus. These data suggest that food consumption induced by 2-deoxyglucose treatment can be antagonized by anorectic drugs acting at recognition sites present in several hypothalamic nuclei, while deprivation-induced feeding acts through different receptor mechanisms which may be specific to the perifornical region of the lateral hypothalamus.

Regulation of [3H]mazindol binding to subhypothalamic areas: Involvement in glucoprivic feeding

The distribution of low-affinity sodium-sensitive binding sites of [3H]mazindol were studied in rat hypothalamic nuclei. Using microdissection methods, it was demonstrated that the highest level of [3H]mazindol binding is localized to the paraventricular nucleus (PVN) and the lowest binding is observed in the lateral hypothalamus. Following food deprivation, a significant decrease in [3H]mazindol binding in the PVN and ventromedial hypothalamus (VHM) were observed. Refeeding food-deprived rats resulted in restoration of the level of binding in the PVN, and this was correlated with changes in blood glucose levels. Thus, changes in the binding of [3H]mazindol in the PVN may reflect local changes in glucose levels. In related studies, the involvement of the PVN in the regulation of food deprivation or 2-deoxyglucose (2-DG)-induced food intake was studied. Application of amphetamine (20 micrograms) into the PVN had no effect on food deprivation induced feeding, but significantly inhibited 2-DG induced (glucoprivic) feeding. The PVN may play an important role in the glucostatic regulation of feeding and in mediating the anorectic action of amphetamine and related anorectic drugs on glucoprivic feeding.

The effect of serotonergic and dopaminergic lesions on sodium-sensitive [3H]mazindol binding in rat hypothalamus and corpus striatum

The effects of intracerebroventricular administration of 6-hydroxydopamine (6-OHDA) and 5,7-dihydroxytryptamine (5,7-DHT) on sodium-sensitive [3H]mazindol binding were investigated in the rat hypothalamus and corpus striatum. In the hypothalamus, specific [3H]mazindol binding was inhibited by low concentrations of sodium and stimulated by high-sodium concentrations, whereas in the corpus striatum, only a sodium-dependent stimulation of [3H]mazindol binding was observed. Lesions with 6-OHDA significantly reduced sodium-dependent [3H]mazindol binding in the corpus striatum, but had no effect on the binding of [3H]mazindol in the absence of sodium. Lesions of serotonergic neurons with 5,7-DHT, however, had no effect on [3H]mazindol binding in the striatum, but resulted in a significant increase in the number of [3H]mazindol binding sites in the hypothalamus. These data suggest that [3H]mazindol may bind to two anatomically distinct binding sites, one that is stimulated and the other inhibited by sodium. The sodium-stimulated binding sites appear to be located on dopaminergic terminals in the striatum, and in the hypothalamus, the sodium-inhibited sites appear to be regulated by serotonergic neuronal activity.