Receptors In Rat Brain Membranes Research Articles

1-Methyl-beta-carboline (harmane), a potent endogenous inhibitor of benzodiazepine receptor binding.

The interaction of several beta-carbolines with specific [3H]-flunitrazepam binding to benzodiazepine receptors in rat brain membranes was investigated. Out of the investigated compounds, harmane and norharmane were the most potent inhibitors of specific [3H]-flunitrazepam binding, with IC50-values in the micromolar range. All other derivatives, including harmine, harmaline, and several tetrahydroderivatives were at least ten times less potent. Harmane has been previously found in rat brain and human urine, so it is the most potent endogenous inhibitor of specific [3H]-flunitrazepam binding known so far, with a several fold higher affinity for the benzodiazepine receptor than inosine and hypoxanthine. Thus, we suggest that harmane or other related beta-carbolines could be potential candidates as endogenous ligands of the benzodiazepine receptor.

Central nervous system stimulating and depressing drugs as possible ligands of the benzodiazepine receptor

The interaction of several central nervous system stimulating and depressing drugs with specific [ 3H]flunitrazepam binding to benzodiazepine receptors in rat brain membranes was investigated. Most of the investigated intravenous anesthetics, hypnotics, anticonvulsives, and analeptics displace specific [ 3H]f1unitrazepam binding at high concentrations, but only the affinities of glutethimide and fominobene for the benzodiazepine receptor (IC 50-values of 42 and 1 μM respectively) are high enough to suggest some pharmacological relevance of the observed interaction with the benzodiazepine receptor. Since the pharmacological properties of the respiratory stimulant fominobene suggest that this drug has rather antagonistic than agonistic properties at the benzodiazepine receptor, fominobene may become useful for elucidating the molecular of action of the benzodiazepines.

Electrophysiological evidence that GABA potentiation in vivo correlates with benzodiazepine receptor affinity

Since the inhibition of cortical neurons evoked by electrical stimulation of the surface of the cerebral cortex is believed to be mediated by endogenous GABA, the effects of five benzodiazepines on this inhibition were studied in cats anesthetized with fluothane or methoxyflurane. The benzodiazepines used were Ro 217ndash;3981, flurazepam, chlordiazepoxide, medazepam and clozapine, because their water solubility allows their microiontophoresis. The degree of inhibition of single cortical neurons induced to fire submaximally by Na L-glutamate was measured from peristimulus histograms generated by the extracellularly recorded spikes. Both the percentage of GABA potentiation by equal iontophoretic doses of the five benzodiazepines, as well as the iontophoretic doses of the benzodiazepines necessary to induce the same degree of GABA-potentiation, agreed well with the relative affinities of these drugs for the benzodiazepine receptor in rat brain membranes in vitro as reported by Braestrup and Squires [2], which are known to correlate well with several pharmacological tests predictive of anxiolytic activity in man.

Tricyclic antidepressant radioreceptor assay

A radioreceptor assay for tricyclic antideressants described here is based on the ability of these drugs to compete with [ 3H]-3-quinuclidinyl benzatile ( 3H-QNB) for binding to muscarinic cholinergic receptors in rat brain membranes. The assay is sensitive, in that it can detect, for example, 2 ng/ml nortriptyline in plasma. Seven plasma samples from depressed patients treated with notriptyline were assayed with the radioreceptor and gas liquid chromatographic methods, and the results for these two methods were almost identical. This assay should be used continously, if at all, in patients treated with other drugs that have potent anticholinergic effects.

Properties of GABA transport puring development in mouse brain

Since the inhibition of cortical neurons evoked by electrical stimulation of the surface of the cerebral cortex is believed to be mediated by endogenous GABA, the effects of five benzodiazepines on this inhibition were studied in cats anesthetized with fluothane or methoxyflurane. The benzodiazepines used were Ro 217ndash;3981, flurazepam, chlordiazepoxide, medazepam and clozapine, because their water solubility allows their microiontophoresis. The degree of inhibition of single cortical neurons induced to fire submaximally by Na L-glutamate was measured from peristimulus histograms generated by the extracellularly recorded spikes. Both the percentage of GABA potentiation by equal iontophoretic doses of the five benzodiazepines, as well as the iontophoretic doses of the benzodiazepines necessary to induce the same degree of GABA-potentiation, agreed well with the relative affinities of these drugs for the benzodiazepine receptor in rat brain membranes in vitro as reported by Braestrup and Squires [2], which are known to correlate well with several pharmacological tests predictive of anxiolytic activity in man.

Interaction of phencyclidine ("angel dust") with a specific receptor in rat brain membranes.

[3H]Phencyclidine binds to synaptic membranes from rat brain in a saturable, reversible, and selective fashion, with a dissociation constant Kd of 0.25 microM and a maximal binding capacity of 2.4 pmol/mg of membrane protein--i.e., 250 pmol/g of brain. The binding activity is concentrated in synaptosomal fractions, is higher in cerebral cortex and corpus striatum than in other parts of the rat brain, and is not detectable in the spinal cord. Only molecules of the phencyclidine series and ketamine are able to bind to the phencyclidine receptor. [3H]Phencyclidine bound to its receptor is not displaced by the classical neurotransmitters or neuromodulators. There is a good correlation between the apparent affinities of a series of phencyclidine analogs for the phencyclidine receptor and the pharacological activities of these analogs as measured by the rotarod assay.

Pharmacological characterization of benzodiazepine receptors in the brain

Receptors in rat brain membranes which specifically bind 3H-diazepam were characterized pharmacologically using reference substances representing several pharmacological classes of drugs. Of 28 benzodiazepines tested, several “classical” ones (diazepam, clonazepam, lorazepam, oxazepam, nitrazepam, flurazepam, bromazepam and chlorazepate) with known clinical efficacy, as well as three newer “triazolo” benzodiazepines (estazolam, U 35,005, U 31,957), one new “imidazolo” benzodiazepine, U 31,219, and one new 2-carbamoylmethylene-benzodiazepine, displaced 3H-diazepam binding at low concentrations (K i = 1–60 nM). For these benzodiazepines there was a statistically significant correlation between K i values for displacement and ED 50 (or MED) values in several pharmacological tests predictive of anxiolytic activity in man. More than 100 nonbenzodiazepines, representing 22 distinct pharmacological classes as well as 14 presumed neurotransmitters in the CNS, including 4 peptides, were much weaker as 3H-diazepam displacers (K i > 0.1 nM). These results that in vitro 3H-diazepam binding represents the physiologically relevant binding to hitherto unknown receptors in the CNS.

The opiate receptor binding interactions of 3H-methionine enkephalin, an opioid peptide

3H-Methionine enkephalin binds stereospecifically with high affinity to opiate receptors in rat brain membranes. Equilibrium experiments indicate two distinct dissociation constants with K D values of 1.8 and 5.8 nM respectively. 3H-Methionine enkephalin associates and dissociates from the opiate receptor with 8–10 fold slower kinetics that 3H-opiates. Though several opiates have similar affinities for sites labeled by 3H-methionine enkephalin, 3H-dihydromorphine and 3H-naloxone, some opiates such as morphine, dihydromorphine and oxymorphone are only one tenth as potent in competing for 3H-methionine enkephalin as for 3H-dihydromorphine and 3H-naloxone binding. As with other opiate agonists, 5–10 mM sodium selectively decreases the binding of 3H-methionine enkephalin. At 26°C, 0.1–1.0 mM managanese but not magnesium or calcium increases the binding of 3H-methionine enkephalin, while at 0°C managanese decreases the binding of methionine enkephalin.