3H-DHE Binding Research Articles

Platelet adrenoceptors and prostaglandin responses in depressed patients

Platelet α 2-adrenergic receptor binding and prostaglandin responsivity were measured in depressed patients. Depressed patients had significantly higher platelet 3H-dihydroergocryptine ( 3H-DHE) binding values than controls. Depressed patients also showed significantly reduced prostaglandin E 1- stimulated cyclic adenosine 3',5'-monophosphate (cAMP) production and significantly decreased % inhibition of cAMP production by norepinephrine. These results support the suggestion that there may be a dissociation between α 2-adrenergic receptor binding and responsivity in depression. There were no significant correlations between platelet adrenergic variables and other indices of noradrenergic function. However, there was a significant correlation between 3H-DHE binding values and basal plasma levels of cortisol.

Noradrenergic and neuroradiological abnormalities in tardive dyskinesia

Previous studies suggest that catecholaminergic overactivity and structural brain damage may contribute to the pathogenesis of tardive dyskinesia (TD). Although dopaminergic (DA) mechanisms, specifically postsynaptic receptor supersensitivity, have been extensively studied, equally plausible noradrenergic (NE) changes have been all but ignored. Likewise, the interaction of neurochemical and neuroradiological abnormalities has received little attention. Over the past 6 years, 111 inpatients were studied with a battery of neurological, behavioral, biochemical, and neuroradiological measures. Forty-one patients met specific diagnostic criteria for TD, based in part on global ratings on the Abnormal Involuntary Movement Scale. Subgroups of patients were also evaluated with the Brief Psychiatric Rating Scale and were assayed for plasma dopamine-β-hydroxylase (DBH) activity, platelet 3H-dihydroergocryptine ( 3H-DHE)-alpha 2 adrenergic receptor binding, lumbar cerebrospinal fluid (CSF) monoamines and metabolites [NE, 3-methoxy-4-hydroxyphenylglycol (MHPG), DA-sulfate, homovanillic acid (HVA), dihydroxyphenylacetic acid (DOPAC)], and CT scan indices of brain atrophy, including ventricle/brain ratio (VBR), bifrontal/bicaudate ratio, and cortical atrophy. All patients were studied in the steady state, primarily when free of neuroleptics. Patients with TD had significantly greater DBH activity than those without TD. In addition, 3H-DHE binding and CSF NE were significantly correlated with the severity of TD when present. Finally, TD patients with low DBH activities (below the mean) had significantly larger ventricles than non-TD patients with low DBH activities. Other data suggested that subcortical, rather than cortical, atrophy was more likely to be responsible for the larger VBR in the low DBH TD group. These results suggest an association of NE overactivity and TD in a portion of patients. Moreover, the presence of neuroradiological abnormalities in TD patients with low DBH activity underscores the contribution of heterogeneous factors to the pathogenesis of this disorder and may provide one possible explanation for the discrepant biochemical findings in TD reported by earlier investigators.

Differential effects of nifedipine, verapamil, and diltiazem on noradrenaline-induced contractions, adrenergic transmitter release, and alpha-adrenoceptor binding in the female rabbit urethra.

In order to study differences in action between "Ca2+-entry blockers" on smooth muscle and peripheral nerves, the effects of nifedipine, verapamil, and diltiazem on noradrenaline (NA)-induced contractions and electrically evoked release of 3H-NA were investigated in the female rabbit urethra. In addition, possible influences of Ca2+-entry blockers on alpha-adrenoceptors were studied with radioligand binding technique. Exposure to Ca2+-free medium completely abolished the contractile response to 1 microM NA in the rabbit urethra, indicating that the contraction was entirely dependent on influx of extracellular Ca2+. The Ca2+-entry blockers inhibited the NA-induced contractions in the following order of potency: nifedipine greater than verapamil approximately equal to diltiazem. In contrast to nifedipine and diltiazem, which produced a maximum inhibition of between 50 and 60%, verapamil was able to abolish the contractile responses to NA. The electrically evoked efflux of 3H-NA was decreased by diltiazem and increased by verapamil, whereas nifedipine failed to alter the 3H-NA efflux. Only verapamil was effective in inhibiting specific 3H-DHE binding to a crude membrane preparation of the rabbit bladder base and urethra, and the inhibition appeared to be of the competitive type. It is suggested that the effects of verapamil on electrically evoked efflux of 3H-NA and on NA-induced contractions can be partly explained by blockade of pre- and post-junctional alpha-adrenoceptors. The failure of nifedipine and diltiazem to abolish the NA-induced contraction might indicate the existence of different Ca2+-entry pathways in urethral smooth muscle.

Influence of antimuscarinics on alpha-adrenoceptors in the female rabbit urethra.

The effects of the tertiary amine atropine and its structural analogues homatropine and scopolamine, as well as the quarternary amine emeprone, were evaluated on noradrenaline (NA)-induced contractions of isolated female rabbit urethral ring preparations. In addition, the abilities of these antimuscarinics to inhibit 3H-dihydro-alpha-ergocryptine (3H-DHE) binding to alpha-adrenoceptors were studied on a crude membrane preparation from the female rabbit bladder base and urethra. Atropine and homatropine depressed the NA-induced contractions in a concentration-dependent way, whereas this was not seen with scopolamine. Emeprone 10(-5) and 10(-4) M augmented the contractions, an effect possibly attributable to a NA-uptake blocking effect. All antimuscarinics displaced specific 3H-DHE binding, the order of potency being atropine greater than homatropine greater than emeprone greater than scopolamine. In general a good correlation was seen between the binding and mechanical activity studies for atropine, homatropine and scopolamine, while this was not found for emeprone. It is concluded that alpha-adrenoceptor blockade by atropine can be observed at concentrations exceeding 10(-7) M. Scopolamine, showing alpha-adrenoceptor blocking properties only in high concentrations, may be used as an alternative for blockade of muscarinic cholinoceptors.

Platelet alpha-adrenergic binding and biochemical responsiveness in depressed patients and controls

In a study of platelet α 2-adrenergic receptor number in depressed patients, binding of tritiated dihydroergocriptine ( 3H-DHE) to platelet membranes was measured in 23 depressed patients and 51 controls. To examine the functional responsiveness of the platelet α 2-adrenergic receptor, basal cyclic adenosine 3',5'- monophosphate (cAMP) production, prostaglandin E 1 (PGE 1) stimulation of cAMP production, and norepinephrine (NE) inhibition of PGE 1-stimulated cAMP production were measured in 23 depressed patients and 53 control subjects. Finally, plasma NE concentration was measured in 20 patients to explore the possible relationship between this endogenous agonist and platelet α 2-adrenergic receptor function. 3H-DHE binding to platelet membranes was significantly increased in the depressed patients compared to control subjects. Both the PGE 1-stimulated cAMP response and the inhibition of this response by NE were significantly reduced in the depressed patients compared to the control subjects. Thus, an apparent dissociation between α 2-adrenergic receptor binding and functional responsiveness was observed. Plasma NE concentrations were neither significantly different in the depressed patients than in the controls nor correlated with any of the measures of cAMP responsiveness. They were, however, significantly negatively correlated with 3H-DHE binding in depressed patients with adequate PGE 1 stimulation of cAMP production.

Demonstration of alpha-adrenoceptors in the rabbit bladder base and urethra with 3H-dihydroergocryptine ligand binding.

The purpose of this work was to demonstrate the presence of alpha-adrenoceptors in a crude membrane preparation made from rabbit bladder base and urethra. This was achieved by radioligand binding studies, using 3H-dihydro-alpha-ergocryptine (3H-DHE) as the radioligand. The specific binding, i.e. the binding that could be inhibited by 10(-5) M phentolamine, was saturable with 73 fmol 3H-DHE bound per mg membrane protein. Binding was at steady state after 60 min., and reversible. Rate constants for association and dissociation were 3 X 10(7) M-1 min.-1, and 2 X 10(-2) min.-1, respectively. A number of compounds were tested for their abilities to compete with 3H-DHE for the binding sites. The relative affinity of some adrenoceptor agonists was: (-)-adrenaline greater than (-)-noradrenaline much greater than (+/-)-isoprenaline. Stereoselectivity was shown, since (-)-noradrenaline had 42 times higher affinity than (+)-noradrenaline. Adrenoceptor antagonists inhibited 3H-DHE binding in the following order of potency: DHE greater than phentolamine much greater than (+/-)-propranolol. The dissociation constant, KD, for DHE to the binding sites was estimated in three different ways. The constants were derived from saturation, competition, and kinetic studies, and gave KD values of 1.1, 1.4 and 0.7 nM, respectively. The results suggest that alpha-adrenoceptors were labelled by 3H-DHE in the tissue homogenates.

Clinical studies of monoamine receptors in the affective disorders and receptor changes with antidepressant treatment

1. 1. Pre-clinical and clinical studies suggest that the responsiveness of monoamine and cholinergic receptors may be altered in the affective disorders and that antidepressants may modify the sensitivity of these receptors. 2. 2. The growth hormone response to clonidine is reduced in depressed patients compared to controls according to several independent studies, suggesting that post-synaptic α 2-adrenergic receptors may be less responsive in depressed patients. 3. 3. The cortisol response to clonidine is enhanced in depressed patients compared to controls in our study raising the possibility that cortisol hypersecretion in depressed patients may be related to noradrenergic dysfunction. 4. 4. The hypotensive response to clonidine is blunted in patients on chronic antidepressant treatment with either clorgyline or desipramine suggesting that pre-synaptic α 2-adrenergic receptors may subsensitize with chronic antidepressant treatment. 5. 5. The prolactin increase in response to fenfluramine is less in depressed patients compared to controls suggesting decreased functional activity of the serotonergic system in depression. 6. 6. Platelet α 2-adrenergic receptor number as measured by tritiated dihydroergocriptine ( 3H-DHE) binding is increased in depressed patients compared to controls, while cyclic 3'–5' adenosine monophosphate (cAMP) production in response to prostaglandin e 1 (PGE 1) and norepinephrine (NE) inhibition of PGE 1-stimulated cAMP production are reduced in the platelets of depressed patients. Thus, it is not clear that increased 3h-dhe binding reflects increased functional responsiveness and might in fact be compensatory to decreases in functional responses of α 2-adrenergic receptors.

Adrenergic regulation on estrogen action and cyclic GMP.

Adrenergic regulation and significance of cyclic GMP on estrogen action in ovariectomized rat uterus were studied in vivo and in vitro. Estradiol-176 (i.p., 1 μg/100g) increased in uterine cyclic GMP significantly (peak at 120 min). These increases were abolished by the pretreatment with phentolamine, but not atropine and propranolol. 3H-DHE binding sites in the uterus treated with estradiol-17β 90 min before were increased significantly. Secondly, the adrenergic regulation of estradiol (5 × 10−12 M) uptake in rat uterus was investigated in normal or Ca2+-free Tyrode’s solution at 30°C. Cyclic GMP content was also investigated under these conditions. L-noradrenaline (4 × 10−7 M) increased 3H-estradiol uptake in both cytosol and nuclear fractions Ca2+-dependently. These increases were abolished in the presence of phentolamine but not propranolol. Increase in uterine cyclic GMP contents was shown under the condition in which 3H-estradiol uptake was stimulated. These results suggested that increase in uterine cyclic GMP depended on increase in α-stimulation, and the estradiol influx involves Ca2+-dependent adrenergic α-regulation in addition to non-specific diffusion; the increase in uterine cyclic GMP was cinsidered to be dependent on the formation of estrogen-receptor complex. The changes in uterine cyclic GMP may serve as an index of estrogen action.

Amitriptyline binding to specific alpha-adrenergic receptors in vitro

Amitriptyline was found to compete with ( 3H) dihydroergocryptine ( 3H-DHE), a potent alpha adrenergic antagonist, for specific alpha-adrenergic binding sites in rabbit uterine membranes preparation. Amitriptyline and adrenergic agonists compete for ( 3H) DHE binding sites in the following order of potency : (−) epinephrine > (−) nor-epinephrine > amitriptyline > ethylphenylephrine. Amitriptyline and alpha and beta antagonists compete in the order : phentolamine ⪖ phenoxybenzamine > amitriptyline > propranolol. Based on 50 per cent inhibition values from binding competition curves, the affinity of amitriptyline for alpha-adrenergic sites was 83-fold lower than phentolamine, but only ten fold lower than epinephrine. These data are consistent with the hypothesis that the hypotensive action of amitriptyline results at least in part from alpha-adrenergic blockade.

Action of detergents on 3H-dihydroergokriptine binding and localization of α-adrenoceptors in synaptosomal membranes

3H-dihydroergokriptine ( 3HDHE) binding was carried out in synaptosomal membranes from basal ganglia of the cat. A single type of binding site with Kd 3.7 nM, Hill number = 0.95 and B max = 1000 pmol/g protein was found. 3HDHE bound to α-adrenoceptors and not to serotonin or dopamine receptors. At very low concentrations, some detergents enhanced binding, but at higher concentrations all those used (Triton X-100, Nonidet P-40, deoxycholate and digitonin), inhibited the binding of 3HDHE. After binding to the membrane protein, the 3HDHE-receptor complex was stable to the action of Triton X-100. At concentrations of Triton X-100 (0.1–0.2%), in which only the presynaptic membrane disintegrated, the 3HDHE specific radioactivity was reduced. With a more drastic treatment that disintegrated the postsynaptic membrane, 3HDHE binding was further reduced. These result suggest that α-adrenergic receptors may be localized at both the pre- and postsynaptic membranes of central synapses.

Platelet Function in Essential Thrombocythemia

Platelets from two patients with essential thrombocythemia failed to aggregate or release serotonin in response to concentrations of epinephrine that aggregated platelets from normal controls. Therefore, we studied their alpha-adrenergic receptors, using 3H-dihydroergocryptine (3H-DHE), an alpha-adrenergic antagonist. These platelets contained an average (mean +/- S.E.) of 210 +/- 18 and 227 +/- 27 3H-DHE binding sites per platelet--less than half that found on control platelets, 464 +/- 37 (P less than 0.01). In contrast, platelets from two other patients with essential thrombocythemia responded to epinephrine and contained a normal number of 3H-DHE sites. Platelets in essential thrombocythemia demonstrated normal kinetics of 3H-DHE binding and normal affinities for 3H-DHE and for epinephrine. When control platelets were preincubated with a half-saturating concentration of 3H-DHE, there was a diminution of epinephrine-induced platelet function comparable to that seen in essential thrombocythemia. Thus, a deficiency of alpha-adrenergic receptors may account for diminished functional responsiveness of platelets to epinephrine in some patients with essential thrombocythemia.

Dihydroergotamine binding to rat brain membranes

3H-dihydroergotamine, which is used clinically to treat orthostatic hypotension and migraine, binds saturably, reversibly and with high affinity (K D = 0.2 nM) to rat brain membranes. The binding is time, temperature and pH dependent and is highest in the hippocampus and the corpus striatum. Serotonin was the only neurotransmitter tested capable of inhibiting 3H-DHE binding.