Potential development of tolerance to and dependence on benzodiazepine tranquilizers often limit their use for long-term treatment of epilepsy, anxiety and insomnia. Current developments in benzodiazepine receptor pharmacology, i.e. the advent of partial agonists and receptor subtype specific agonists (Fig. 1), however, might eventually overcome these limitations, thus greatly improving therapeutic prospects. The present study demonstrates that subchronic administration of alprazolam (a high-efficacy agonist) results in strong withdrawal reactions upon injection of a benzodiazepine receptor antagonist in mice and monkeys. The partial agonist bretazenil, as well as the benzodiazepine receptor type 1-preferring agonist zolpidem, however, are much less prone to producing such reactions. Neurochemical studies showed that subchronic infusion of lorazepam (a high-efficacy agonist), in contrast to betrazenil, led to benzodiazepine receptor downregulation in vivo and reduced potentiation of gamma-aminobutyric acid (GABA)-stimulated chloride flux by diazepam ex vivo. These findings indicate that partial and receptor subtype 1-selective agonists differ from full, non-selective agonists in their liability to induce drug dependence and tolerance upon chronic administration. It is hypothesized that the neurochemical basis of these adaptive phenomena may be receptor downregulation and/or reduced coupling between GABAA receptor/chloride channel gating and benzodiazepine receptor binding.

The translocation of protein kinase C (PKC) from the cytosol to the membrane might be functionally involved in learning and memory. Using [3H]-phorbol 12,13-dibutyrate (3H-PDBu) binding three pools of binding sites could be distinguished in tissue preparations: Pool a comprised the soluble receptors which bound phorbol ester with low affinity in the absence of calcium. Pool b was composed of high-affinity phorbol ester binding sites identified in the soluble fraction upon addition of calcium. Pool c represented stably membrane-bound receptors binding phorbol ester independently of calcium. 3H-PDBu binding was then measured in the cortices and hippocampi of rats trained in an eight-arm radial maze. A progressive training-dependent increase of membrane-bound binding activity with a concomitant decrease in the soluble fraction was detected independent of learning the maze task. These results suggest that it is the experience of an enriched environment by the repeated behavioral stimulation in a maze rather than the acquisition of a memory task that leads to enhanced incorporation of phorbol ester receptors (PKC) into the cell membrane.

The benzodiazepine receptor is an allosteric modulatory site present on most, if not all, gamma-aminobutyric acid A (GABAA) receptor channels (GABAA-R). The benzodiazepine receptor recognizes a large spectrum of compounds from different chemical classes that are grouped together as benzodiazepine receptor ligands--of benzodiazepine and non benzodiazepine structure. The GABAA-R is thought to be a heteropentameric protein complex composed of at least three different classes of subunits, with each subunit comprised of up to six structural variants. Binding of GABA to the extracellular domain of the receptor causes a conformational change that opens the channel pore to anions. A classical benzodiazepine achieves a positive allosteric modulation of the GABA channel gating function by increasing the affinity of the receptor for GABA and, possibly, by facilitating the conformational transition from the closed to the open form (benzodiazepine receptor agonists). Inverse agonists of benzodiazepine receptors cause negative allosteric modulation (a decrease in the GABA activity). Benzodiazepine receptor antagonists bind to the benzodiazepine receptor with little effect on GABAA-R functioning. The intrinsic efficacy of benzodiazepine receptor ligands determines the direction and magnitude of allosteric modulation. Benzodiazepine receptor agonists affect neuronal activity in all major neuronal networks. The classical pharmacological profile of benzodiazepine receptor agonists consists of anxiolytic, anticonvulsant, sedative, and myorelaxant activities. Partial agonists of benzodiazepine receptors conserve anxiolytic and anticonvulsant activity, with greatly reduced sedation and muscle relaxation. They promise to present therapeutic advantages, in particular for long term use. In initial studies. they have produced fewer side-effects and showed reduced tolerance development and physical dependence liability.(ABSTRACT TRUNCATED AT 250 WORDS)

Intrinsic efficacy is the inherent ability of a ligand to induce the conformational change of its receptor that is required to transduce the event of signal recognition into a physiologically relevant response. Relating fractional receptor occupancy to fractional effect is an indirect but reliable way to assess relative intrinsic efficacy. The receptor studied was the benzodiazepine receptor (BZR), a modulatory site on the gamma-aminobutyric acidA (GABAA) receptor-chloride channel. The relationship between fractional BZR occupancy, as assessed by inhibition of [3H]flumazenil binding, and potentiation of GABA-stimulated 36Cl- influx into membrane vesicles of rat cerebral cortex was evaluated for four ligands under identical experimental conditions. Triazolam and the quinolizinone Ro 19-8022 potentiated the effect of GABA maximally by nearly 50%, diazepam by about 40% and bretazenil by approximately 20%. Potentiation of GABA-stimulated 36Cl- flux by 25% was observed at about 35% BZR occupancy for diazepam, about 45% for triazolam and about 95% for Ro 19-8022. Bretazenil did not produce 25% potentiation even at receptor saturation. Although the curves relating fractional BZR occupancy to GABA potentiation were hyperbolic and nearly superimposable for triazolam and diazepam, those for Ro 19-8022 and bretazenil displayed parabolic characteristics by inducing an effect only at very high BZR occupancy, reflecting the partial agonistic profile of the latter two compounds. The rank order of relative intrinsic efficacy determined in this study was: triazolam congruent to diazepam much greater than Ro 19-8022 greater than bretazenil.

Agonists at the benzodiazepine receptor (BZR) produce their effects through potentiation of the inhibitory alpha-aminobutyric acid-mediated neurotransmission in the central nervous system via positive allosteric modulation of the gamma-aminobutyric acidA receptor. Agonists with high intrinsic efficacy are anticonvulsant, anxiolytic, muscle relaxant and sedative, whereas agonists with low intrinsic efficacy (partial agonists) are predominantly anticonvulsant and anxiolytic, but antagonize muscle relaxant and sedative effects of full agonists. The four BZR ligands triazolam, diazepam, Ro 19-8022 (a benzoquinolizinone) and bretazenil (Ro 16-6028, an imidazobenzodiazepinone) were pharmacologically characterized in various neurological and behavioral paradigms in mice: two anticonvulsant tests (prevention of audiogenic and pentylenetetrazol-induced seizures), a conflict test which reveals both anxiolytic and sedative properties and two tests which mainly measure motor impairment (rotarod and horizontal wire test). Although triazolam and diazepam elicited an effect in all tests, Ro 19-8022 and bretazenil exhibited anticonvulsant and anxiolytic properties, but virtually failed to induce motor impairment and severe sedation. In separate experiments, fractional BZR occupancy in vivo was assessed by inhibition of [3H]flumazenil binding and correlated with the pharmacological effects. Although diazepam and triazolam produced effects beginning at low to intermediate fractional BZR occupancy, Ro 19-8022 and bretazenil required a higher BZR occupancy to do so, in accordance with their partial agonistic character. With the two full agonists, anticonvulsant and anticonflict activities were elicited at a lower fractional BZR occupancy than muscle relaxant and sedative effects.(ABSTRACT TRUNCATED AT 250 WORDS)

In the present study the effect of acute administration of nebivolol (NBV) on increasing current electroshock (ICES) test, PTZ (pentylenetetrazole) induced seizures, spatial working memory and locomotor activity on rotarod in mice alone as well as in combination with gabapentin (GBP) were evaluated.The anticonvulsant effects of the drugs were measured using ICES and PTZ model whereas cognitive behavior was measured by the spontaneous alternation behavior and rotarod test. The biochemical estimation was done by measuring the lipid peroxidation and reduced glutathione (GSH).Significant increased seizure threshold in ICES test and latency to clonic jerks in PTZ induced seizures test was found by NBV alone as well in combination with GBP. There was no effect on spatial working memory and locomotor activity. In addition NBV in combination with GBP significantly decreased the level of the lipid peroxidation and increased the level of GSH in brain.The Study showed that NBV potentiates the anticonvulsive activity of GBP, which can be useful for the treatment of epilepsy in patients with hypertension.

Benzodiazepines in clinical use have a range of pharmacological activities. Some, e.g. sedation, tolerance and addiction, are not welcome. Undesirable side-effects of drugs are often controlled by developing compounds that bind more selectively to one particular receptor subtype. An alternative approach, discussed here by Willy Haefely and colleagues, is the development of partial agonists which exploit regional differences in receptor reserve to tease apart biological responses. Partial agonists for the benzodiazepine modulatory site on the GABAA complex have been developed and their pharmacological profiles can be interpreted to suggest that neurons mediating anticonvulsant and anti-anxiety effects do indeed have a higher receptor reserve than neurons mediating other unwanted effects. This suggests that benzodiazepine receptor partial agonists may have important therapeutic potential.

Tracheobronchial colonization by Gram-negative bacteria is common in mechanically ventilated patients. Pseudomonas sp are commonly isolated from the lower airways. We hypothesized that Pseudomonas sp would preferentially colonize the lower airway and would be more common in patients with poor nutritional status. We serially collected 75 pairs of upper and lower respiratory tract cultures from 14 patients treated with mechanical ventilation for at least one week, examined patterns of airway colonization and routes of bacterial entry for Pseudomonas sp and other enteric Gram-negative bacteria (EGNB), and related these findings to host-associated factors, including nutritional status. Pseudomonas sp were the most common species isolates taken from the lower airway, found in nine of 14 patients and in 41.3 percent of all cultures. In contrast to other EGNB, Pseudomonas sp were found significantly (p less than or equal to 0.05) more often in the tracheobronchial tree (31 of 75 cultures) than in the oropharynx (18 of 75 cultures). Primary colonization of the lower airway by Pseudomonas sp was found in four patients, while other EGNB never followed this pattern when subjects were studied with cultures taken every third day. A host-related factor related to lower airway colonization by Pseudomonas species was poor nutritional status, assessed by a multifactorial index (p less than or equal to 0.01). We conclude that in mechanically ventilated patients, Pseudomonas sp colonize the lower airway in a different pattern and by a different route from those of other EGNB. The findings that Pseudomonas sp preferentially colonize the tracheobronchial tree may be important for the design of strategies to prevent airway colonization. The recognition that poor nutritional status, a potentially modifiable host-related factor, favors lower airway growth of Pseudomonas sp suggests one direction for future infection-control efforts.

The imidazobenzodiazepinone derivative Ro 15-4513 has the activity profile of a partial inverse (low efficacy) agonist at the benzodiazepine receptor (BZR). It reverses central nervous depressant effects of diazepam, and, in part, of phenobarbitone and ethanol in mice, rats and cats in behavioural, electrophysiological, and neurochemical paradigms. The interaction of Ro 15-4513 with barbiturates and ethanol is due to its inverse agonistic (negative allosteric modulatory) property at the BZR, as it was reversed by the selective BZR blocker flumazenil (Ro 15-1788). In the present experiment situations, other BZR partial inverse agonists in subconvulsant or overt convulsant doses were less effective against ethanol effects than Ro 15-4513. Possible mechanisms for this differential activity of BZR inverse agonists are discussed.

GABA-gated chloride channels in the central nervous system contain a regulatory site, the benzodiazepine receptor, through which drugs can modulate the efficiency of GABAergic synaptic transmission and thereby affect the degree of anxiety, muscle tension, vigilance and convulsions. The biochemical analysis of the purified receptor complex with monoclonal antibodies shows a heterooligomeric composition of two glycosylated subunits (alpha, beta). The immunoprecipitated complex contains the binding sites for GABA, benzodiazepines and the convulsant TBPS. The receptor complex was located, immunocytochemically, in synapses of brain regions rich in GABAergic nerve terminals.

Experiments using an electrically biased limiter have been performed on the Texas Experimental Tokamak (TEXT). A small movable limiter is inserted past the main poloidal ring limiter (which is electrically connected to the vacuum vessel) and biased at VLim with respect to it. The floating potential, plasma potential and shear layer position can be controlled. With ¦VLim¦⩾ 50 V the plasma density increases. For VLim < 0 this is primarily a response to an increased particle confinement time, i.e. a decreased outward particle flux at the plasma edge. Instrinsic impurity line behavior is consistent with a reduction of the outward flux at the edge, while impurity laser ablation demonstrates no transport changes in the plasma interior. With VLim > 0 the results obtained are inconclusive. Variation of VLim changes the electrostatic turbulence which may explain the observed total flux changes.

Monoclonal antibodies, raised against a purified GABAA/benzodiazepine receptor complex from bovine cerebral cortex, have been used to visualize the cellular and subcellular distribution of receptorlike immunoreactivity in the rat CNS, cat spinal cord, and bovine and postmortem human brain. Two different antibodies have been used for these studies; bd-17 recognizes the beta-subunit (Mr 55 kDa) in all the species tested, whereas bd-24 recognizes the alpha-subunit (Mr 50 kDa) of bovine and human but not rat and cat tissues. In bovine and human brain, both antibodies produced very similar staining patterns, indicating a homogeneous receptor composition, at least in the brain areas investigated. The general distribution and density of receptor antigenic sites in all tissues studied were very similar to that of benzodiazepine binding sites radiolabeled with 3H-Ro 15–1788 and of glutamate decarboxylase (GAD)-stained nerve terminals. The results demonstrate a very high receptor density (around neuronal cell bodies and processes or less discretely distributed) in the rat olfactory bulbs, cerebral cortex, ventral pallidum, islands of Calleja, globus pallidus, hippocampus, dentate gyrus, substantia nigra, geniculate nuclei, inferior colliculus, cerebellum, reticular formation, spinal cord, and retina. In contrast, no receptors could be detected in white matter, pineal, pituitary, adrenals, and superior cervical ganglia. Only among the cerebellar layers did we observe a conspicuous difference between the staining intensity and the radiolabeling. In bovine and postmortem human brain, e.g., hippocampus, dentate gyrus, cerebral cortex, and substantia nigra, the same close correlation between the immunohistochemical and radiohistochemical findings was observed. At the electron microscopic level, the immune reaction product in the rat substantia nigra and globus pallidus, for example, was localized to pre- and postsynaptic membranes of axodendritic and axosomatic synapses. Whether the presynaptic labeling represents GABA autoreceptors is discussed. In the near future, the monoclonal antibodies will be used in double-labeling experiments with GAD to identify those GABAergic projections that are modulated by benzodiazepine minor tranquillizers. Furthermore, they could also be used, in studies of postmortem human brain, to diagnose receptor dysfunction possibly associated with CNS disorders such as epilepsy.

In mammalian CNS, GABAergic synaptic transmission (GABA = γ-aminobutyric acid) is fundamental to inhibitory feedback and feedforward circuits of projecting neurons and local interneurons. Up to 30% of all synapses in the brain are thought to be GABAergic. The synaptic inhibitory action of GABA is due to the opening of GABA-gated chloride channels, which in turn leads to an increase in the chloride conductance of the subsynaptic membrane. Most frequently, the chloride flux is directed inward, leading to a hyperpolarizing inhibitory postsynaptic potential. In recent years, studies of the GABAergic inhibitory synaptic transmission have unraveled a molecular mechanism by which anxiety, vigilance, muscle tension and the occurrence of convulsions can be regulated.