The time course of the behaviour of GABAA receptors (affinity of muscimol for GABA binding sites and their number) in membranes obtained from five distinct rat brain areas was evaluated in dependence of the decline of the serum concentrations of estradiol and progesterone following hormone deprivation by either ovariectomy (OVX) or ovary- plus adrenalectomy (OVX-ADX) after 1, 2, 3, 5, 14 and 30 days. Diestrus rats served as control. The effect of OVX on the GABA binding sites was due to a decrease of affinity rather than alterations in their density. Conversely, the combined surgery of OVX plus ADX affected both the affinity and the density. Also, the OVX-linked changes of muscimol binding were observed only after five days as compared to control animals, while the initial OVX-ADX associated changes occurred already after one day. OVX-ADX produced significant decreases in the number of binding sites at the shortest times investigated, in frontal cortex (FC) at day one and two and in the medulla oblongata (MED) after three days, which disappeared in the subsequent times studied. Concomitant with the major decrease of steroid hormone serum levels one day after OVX-ADX the FC displayed an increase in affinity by 45%. From the time when steroid serum levels are low the alterations in affinity elicited by both OVX-ADX and OVX only take a similar course, reflected by the following decrease in affinity. In MED an additional adrenalectomy did not significantly modify the OVX linked decrease of affinity, except that the initial effect was already evident two days after OVX-ADX.(ABSTRACT TRUNCATED AT 250 WORDS)

Regional differences in the modulation of [3H] muscimol binding to GABAA receptor complexes by kavapyrones, compounds of the rhizome of the plantPiper methysticum which possess sedative activity, were demonstrated using membrane fractions obtained from target brain centers of kavapyrone action: hippocampus (HIP), amygdala (AMY) and medulla oblongata (MED), and from brain centers outside the main kavapyrone effects as frontal cortex (FC) and cerebellum (CER). The kava extract enhanced the binding of [3H] muscimol in a concentration-dependent manner with maximal potentiation of 358% over control in HIP followed by AMY and MED (main target brain centers). Minimal stimulation was observed in CER followed by FC. In contrast, apart from CER, the potency of kavapyrones was similar in the brain areas investigated with EC50 values ranging between 200 and 300 µM kavapyrones. Scatchard analysis revealed that the observed effects of kavapyrones were due to an increase in the number of binding sites (Bmax), rather than to a change in affinity. At a kavapyrone concentration of 500 µM the order of enhancement in Bmax was HIP=AMY>MED>FC>CER. When kavapyrones are included together with pentobarbital or HPO the two classes of compounds produced a more than additive, i.e., synergetic effect on [3H] muscimol binding. Our findings suggest that one way kavapyrones might mediate sedative effects in vivo is through effects on GABAA receptor binding.

The aim of the present investigation was to answer the question if there exists a relation between the equipment of human gliomas with GABA binding sites and the degree of malignancy of these tumours diagnostically characterized according to WHO classification. The following parameters were assessed: the density, the affinity and the sensitivity to the modulating steroids 3 alpha-hydroxy-5 alpha-pregnane-20-one (3 alpha OH-DHP) and 3 alpha,21dihydroxy-5 alpha-pregnane-20-one (THDOC). Scatchard analysis and non linear computerization revealed that the occurrence of GABA sites was directly related to the degree of tumour malignancy: GABA sites were only detectable in lower malignant gliomas of WHO grade II but not in the very malignant glioblastomas. However, irrespective of the individual density to be detected all glioma GABA sites were sensitive to 3 alpha-hydroxy-5 alpha-pregnane-20-one (3 alpha OH-DHP) and 3 alpha 21dihydroxy-5 alpha-pregnane-20-one (THDOC) without exception. The effects of THDOC were due to increases in the number of binding sites whereas in the presence of 3 alpha OH-DHP a decrease in affinity was noted, additionally. These findings support the view of a normal functional integrity of GABA receptors in gliomas.

The allosteric regulation of specific [3H]-muscimol binding by neuroactive steroids to the GABA-binding sites of membrane fractions prepared from five different brain areas was characterized in order to elucidate if the regionally variable subunit composition of GABAA receptors is reflected in the responsiveness of the GABA binding site to neurosteroid modulatory effects. At a final concentration of 1 microM progesterone and its metabolite 3-alpha-hydroxy-5-alpha-pregnane-20-one (HPO) reduced the affinity in hippocampus (HIP), enhanced it in medulla (MED) and did not affect it in cerebellum (CER). However, there are differences in potency of these two steroids between frontal cortex (FC) and hypothalamus (HYP), since the affinity was enhanced in FC only by progesterone and in HYP only by HPO. While the magnitude of progesterone-induced alterations in affinity were similar in FC, MED and HIP, HPO affected the affinity significantly stronger in HIP than in HYP and MED. Concerning the density of the binding sites progesterone exerted no significant modulatory effect in contrast to HPO which increased the number of binding sites (Bmax) in all five brain areas investigated. However, the enhancements in Bmax were regionally different. The HIP reached the maximal increase of Bmax, followed by FC and MED. The smallest enhancement was found in CER, followed by HYP. Neurosteroidal activity exhibited also THDOC and alphaxalone, the synthetic HPO analogue. A significant different potency of THDOC was found in FC versus CER, whereas alphaxalone did not display regionally different efficacy.(ABSTRACT TRUNCATED AT 250 WORDS)

Regional differences in the effects of neuroactive steroids on the GABAA receptor complex have been demonstrated in vitro (1-3). In order to elucidate if a regionally different modulation of the GABA-binding site also occurs after acute steroidal treatment, in the present study the effects of systemically applied progesterone (10 mg/kg body wt) and its metabolite 3 alpha-hydroxy-5 alpha-pregnane-20-one (3 alpha, 5 alpha-THP) (2 mg/kg body wt) on the specific [3H]muscimol binding to the GABA-binding site was evaluated using membrane fractions prepared from five different brain areas of ovariectomized rats. The receptor density (mumol/kg protein) was not affected in all brain areas investigated by treatment with 3 alpha, 5 alpha-THP but was enhanced selectively in the frontal cortex and hippocampus of progesterone-treated animals. A decrease of affinity was observed in medulla after application of progesterone or 3 alpha, 5 alpha-THP, in contrast to the hypothalamus, which was the single region exhibiting an increase of affinity after steroidal administration. In the frontal cortex and hippocampus only the injection of progesterone reduced the affinity, whereas in cerebellum both steroidal treatments were incapable of affecting the affinity. These regional differences in response to the GABA-binding site to acutely applied progesterone and 3 alpha, 5 alpha-THP are consistent with the existence of heterogeneous populations of GABAA receptor-coupled steroid-binding sites.

3 alpha-Hydroxy-5 alpha-pregnan-20-one (HPO) is a progesterone metabolite which exhibits narcotic properties at high concentrations by interactions with the receptor for gamma-aminobutyric acid (GABA). The present investigation characterized low-dose effects of HPO on GABAA receptor binding, by determining the allosteric properties of HPO on the in vitro binding of 3H-muscimol to membrane fractions from the cerebella of ovariectomized rats. A newly developed method for tissue preparation was used to wash out endogenous ligands interfering with the assay. HPO reduced the affinity of 3H-muscimol to GABAA receptor sites by 52% and enhanced the number of accessible binding sites from 5.5 +/- 0.5 to 7.5 +/- 1.3 pmol/mg protein at subnanomolar (0.1 nM) HPO concentrations. The modulatory effects of HPO on GABAA receptor binding provide evidence that this pregnane steroid might be a physiologically relevant modulator of GABAergic neurotransmission.

The narcotically active progesterone metabolite 3-alpha-hydroxy-5-alpha-pregnan-20-one (HPO) modulates gamma-aminobutyric acid (GABA) neurotransmission by direct actions on the GABA(A)-receptor complex. In the present investigation, the formation of HPO was quantified in man. Twenty-four h urine samples were collected during the night (2300 to 0700 h) and day (0700 to 2300 h) from 11 healthy subjects (31 +/- 5 years) for two consecutive days. The concentration of HPO was measured after enzymatic hydrolysis of conjugated HPO and multiple chromatographic separation steps by gas chromatography-mass spectrometry. The mean excretion rates of HPO were 14.8 +/- 11.8-mu-g/24 h with high inter-individual variations. There were no significant differences in the production of HPO during the night and day. It seemed unlikely that HPO was primarily formed in the adrenals or gonads since the excretion rates of HPO poorly correlated with the formation of 17-ketosteroids or 17-hydroxycorticosteroids that are widely used as indices of adrenal and gonadal steroid production. The data showed for the first time that HPO is produced in mate subjects in concentrations similar to that of classical steroid hormones.

The specific binding of tritiated muscimol to gamma-aminobutyric acid (GABA) receptor sites was studied in distinct brain areas of female rats during different endocrine states. In diestrous rats with intact ovaries the highest receptor densities were found in the cortex (10.24 pmol/mg protein) and the lowest concentrations in the mediobasal hypothalamus (3.29 pmol/mg protein). Four weeks after removal of the ovaries, the number of binding sites was enhanced up to 2.4-fold in all brain areas investigated: the preoptic brain area, mediobasal hypothalamus, corticomedial amygdala, and cerebral cortex. The affinity of the binding sites remained unchanged. Substitution of estradiol and progesterone reduced the number of binding sites to values seen before ovariectomy. The induction of an afternoon surge of LH by estradiol that could be blocked by enhancing the GABAergic tone was accompanied by a distinct reduction in B(max) in the preoptic area in the morning.These results give evidence that ovarian hormones modulate GABAergic neurotransmission by regulation of GABA(A) receptor synthesis or degradation.

Drisseldorf 1 . t, Hbetween the turned on and turned off" form of actin 9 +~ is determmned by the Ca binding to TnC along the actin filament.We shall discuss the essential differences to previous hypotheses and their implications not only for our understanding of muscle contraction and cell motility but also for resulting new pharmacological approaches for modulation of contractile function, especially in myocardium and smooth muscle.Supproted by DFG Br 849/I-2,3.

Journal Article Endocrine regulation of opioid receptors in the female rat brain Get access M Jüptner, M Jüptner Institut für Physiologische Chemie, Universitätsklinikum Essen Search for other works by this author on: Oxford Academic Google Scholar C Hiemke, C Hiemke Psychiatrische Klinik, Universität Mainz Search for other works by this author on: Oxford Academic Google Scholar A Jussofie A Jussofie Institut für Physiologische Chemie, Universitätsklinikum Essen Search for other works by this author on: Oxford Academic Google Scholar Acta Endocrinologica (Norway), Volume 120, Issue 3_Supplement, Jun 1989, Pages S203–S204, https://doi.org/10.1530/acta.0.120S203 Published: 01 June 1989