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Biomedical subjects

L G Miller

Publications and source records attributed to L G Miller.

At least 145 records · Page 8Linked to original sources
PubMed1988

Benzodiazepine binding sites on PC12 cells: modulation by nerve growth factor and forskolin.

PC12 pheochromocytoma cells show increased binding of the peripheral type benzodiazepine Ro 5-4864 after treatment with nerve growth factor (NGF) in membrane preparations. Forskolin, an activator of adenylate cyclase, acts synergistically with NGF to produce further increases in binding, but by itself produces no effect. The increased binding appears to reflect increases in receptor number, since Kd remains unchanged. Binding shows a trend toward increase by day 6 after NGF treatment, and the increase is significant at days 9 and 12. The physiological roles of benzodiazepine binding sites on PC12 cells are unclear. Treatment of the cells with Ro 5-4864 produces no changes in basal or stimulated release of catecholamines, in activity of adenylate cyclase, or in cell proliferation.

Adenine↗
PubMed1988

Modulation of benzodiazepine receptor binding in mouse brain by adrenalectomy and steroid replacement.

Adrenal steroids alter neuronal excitability in the central nervous system (CNS), and evidence from in vitro studies indicates that at least some of these effects are mediated by the GABAergic system. Benzodiazepine receptor binding, among other sites on the GABA complex, has been implicated in steroid-induced alterations in the CNS. To investigate the modulation of benzodiazepine receptor binding by adrenal steroids, we examined receptor binding determined by an in vivo technique in mice after adrenalectomy, hypophysectomy and after replacement with several naturally occurring and synthetic steroids. Benzodiazepine receptor binding was substantially augmented in cortex, hypothalamus, and hippocampus in mice 1 week after adrenalectomy, and these increases appeared to be due to increased receptor number rather than changes in apparent affinity. Similar results in cortex were found after hypophysectomy. Replacement with physiologic, but not lower doses, of corticosterone reversed the changes induced by adrenalectomy. Chronic treatment with deoxycorticosterone also returned binding to control values, but chronic administration with dexamethasone, aldosterone and dihydroprogesterone did not alter binding after adrenalectomy. Adrenalectomy did not alter non-specific binding or GABA concentrations in cortex, and delivery of radioligand did not appear to be affected. These results indicate that adrenal steroids modulate benzodiazepine receptor binding in vivo, perhaps via the CR subtype of corticosteroid receptors. The steroid-benzodiazepine interaction may be especially important in the stress response.

20-alpha-Dihydroprogesterone↗
PubMed1988

'GABA shift' in vivo: enhancement of benzodiazepine binding in vivo by modulation of endogenous GABA.

The enhancement of benzodiazepine binding by gamma-aminobutyric acid (GABA) and its analogues has been described in detail in brain membrane preparations, but results in in vivo preparations such as tissue slices or animals treated with GABA modulators are conflicting. This 'GABA shift' in vitro has been reported for compounds with agonist effects at the benzodiazepine receptor but not for antagonists. We examined the effects of modulators of endogenous GABA on benzodiazepine receptor binding in vivo as determined by specific uptake of the benzodiazepine antagonist [3H]Ro 15-1788. Enhancement of radioligand uptake was observed in cortex, hypothalamus, hippocampus and pons-medulla 4 h after treatment with aminooxyacetic acid (AOAA), in cortex, cerebellum, hypothalamus, hippocampus and pons-medulla 0.5 h after treatment with valproic acid, and in cortex, cerebellum, hypothalamus and hippocampus 6 h after treatment with gamma-vinyl-GABA. GABA concentrations were increased at each of these points, as were synaptosomal GABA concentrations in prior studies. In contrast, no changes in radioligand uptake or GABA concentrations were observed 12 and 24 h after gamma-vinyl-GABA treatment. Increases in binding appeared to be due to increased apparent affinity at the receptor rather than a change in receptor number. These data indicate that binding of a benzodiazepine antagonist undergoes a GABA shift in vivo analogous to that observed with agonists in vitro.

Aminooxyacetic Acid↗
PubMed1988

Kinetics, brain uptake, and receptor binding characteristics of flurazepam and its metabolites.

The benzodiazepine derivative flurazepam (FLZ) is widely used as a hypnotic, but the relative contributions of FLZ and its metabolites desalkylflurazepam (DA-FLZ), hydroxyethylflurazepam (ETOH-FLZ), and flurazepam aldehyde (CHO-FLZ) to overall clinical activity remain uncertain. A single 20 mg/kg dose of FLZ.HCl was administered to mice, with plasma and brain concentrations of FLZ and metabolites determined during 5 h after dosage. Brain and plasma concentrations of FLZ were maximal at 0.5 h after dosage, then declined rapidly in parallel, whereas those of DAFLZ were maximal at 2 h, then declined slowly. Concentrations of ETOH-FLZ, the most polar metabolite, were maximal at 0.5 h, and were undetectable after 3 h. Little CHO-FLZ was detected in either brain or plasma. A single 30-mg oral dose of FLZ.HCl was given to 18 human volunteers, with plasma levels determined over 9 days. FLZ was detected in plasma at low concentrations for no more than 3 h after dosage. ETOH-FLZ concentrations were higher and persisted for 8 h after dosage. CHO-FLZ reached intermediate peak levels and was present longer than FLZ or ETOH-FLZ. In contrast, DA-FLZ achieved the greatest peak concentrations, occurring at 10 h after dosage. Levels declined very slowly, with a mean half-life of 71.4 h, and were still detectable 9 days after FLZ dosage. Plasma free fractions (percent unbound) in mice were 40.3, 51.4, and 25.0% for FLZ, ETOH-FLZ and DA-FLZ, respectively; in humans, values were 17.2, 35.2, and 3.5%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Acute barbiturate administration increases benzodiazepine receptor binding in vivo.

Barbiturates have been reported to augment benzodiazepine receptor affinity in vitro, but their effects in vivo are uncertain. We determined benzodiazepine receptor binding in vivo by specific uptake of [3H]Ro15-1788 after barbiturate administration. Pentobarbital (30 mg/kg) increased receptor binding in cerebral cortex and cerebellum at 30 min after injection, with a peak effect occurring at 1 h after dosage, and a return to control levels at 2 h. Specific binding was increased at 1 h after pentobarbital administration in a dose-dependent fashion (7.5-90 mg/kg). Pentobarbital at doses up to 30 mg/kg failed to alter nonspecific binding, but at doses of 60 mg/kg increases in nonspecific binding were observed. The increases in specific binding observed after barbiturate administration were most likely due to a change in apparent receptor affinity, as determined by administration of varying doses of clonazepam to pentobarbital-treated (30 mg/kg) animals. The order of potency of a series of barbiturates in augmenting benzodiazepine receptor binding in cerebral cortex and cerebellum in vivo was: secobarbital greater than pentobarbital greater than amobarbital greater than phenobarbital greater than barbital. The same relative rank order of potency exists for the anesthetic/hypnotic activity of these barbiturates. These data suggest that barbiturates increase the apparent affinity of benzodiazepine receptors in vivo; unlike their in vitro actions, these alterations can be detected with a receptor antagonist.

Animals↗
PubMed1988

Differential modulation of benzodiazepine receptor binding by ethanol in LS and SS mice.

The LS and SS lines of mice were initially selected based on sedative responses to ethanol, but have been found to differ in response to a variety of hypnotics and anesthetics. These differences do not appear to be due to pharmacokinetic factors and several lines of evidence suggest involvement of the GABAergic system. To examine an important component of this system, the benzodiazepine receptor, we analyzed benzodiazepine receptor binding in vivo in LS and SS mice, and modulation of receptor binding by three interventions known to increase binding in other strains: pentobarbital, defeat stress, and ethanol. Receptor binding was determined by specific uptake of [3H]-Ro15-1788. Receptor binding was increased in cortex and hippocampus of LS mice compared to SS mice, with the increase in cortex most likely due to increased receptor number rather than a change in apparent affinity. Pentobarbital (30 mg/kg IP) induced similar increases in binding in both lines in several brain regions. Defeat stress caused increased binding in several brain regions of both SS and LS mice, with greater binding in cortex of LS mice. In contrast, ethanol at 3 doses (0.5, 1, and 2 g/kg) led to greater increases in binding in SS mice compared to LS mice in most brain regions. None of the interventions altered nonspecific binding. Ethanol concentrations were slightly greater in plasma and brain of LS mice. These results indicate differences in benzodiazepine receptor binding in LS and SS mice, with differential modulation of binding by ethanol but not by pentobarbital or stress. These differences may contribute to differential pharmacodynamic responses in the two lines of mice.

Animals↗
PubMed1988

Low-dose alprazolam augments motor activity in mice.

The triazolobenzodiazepine alprazolam appears to have a unique clinical spectrum, and recent studies indicate unusual binding properties at the benzodiazepine receptor when assessed in vivo at low doses (0.02-0.05 mg/kg). To assess the behavioral activity of alprazolam at low doses, we examined open-field activity after one hour in mice treated with alprazolam, triazolam, and clonazepam. Following triazolam and clonazepam administration, open-field activity decreased in a dose-dependent fashion. In contrast, low doses of alprazolam resulted in an increase in open-field activity, whereas higher doses decreased activity. For all three drugs, activity was linearly related to receptor binding. Pretreatment with a dose of the benzodiazepine antagonist Ro15-1788 sufficient to fully occupy receptors had no effect on open-field activity, but when administered concurrently with alprazolam (0.05 mg/kg) prevented the increase in activity seen with alprazolam alone. Increased open-field motor activity represents a behavioral correlate to the increases in receptor binding seen with low-doses of alprazolam. Changes in activity appear to be mediated at the benzodiazepine receptor, since an antagonist prevents increased activity. These data suggest that the unique clinical effects of alprazolam may be due in part to unusual interactions with the benzodiazepine receptor.

Alprazolam↗
PubMed1988

High-affinity benzodiazepine binding sites on rat peritoneal mast cells and RBL-1 cells: binding characteristics and effects on granule secretion.

High-affinity binding sites for benzodiazepines are present in a wide range of tissues other than brain, and appear to be pharmacologically distinct from central nervous system receptors. Binding sites have been reported on rat peritoneal mast cells, but specificity of binding and functional effects have not been addressed in these cells. Using the specific radioligand [3H]-Ro5-4864, which binds to peripheral but not central binding sites, we characterized binding to rat peritoneal mast cells and the related cell line, RBL-1 cells, and effects on granule secretion. Studies in rat peritoneal mast cells disclosed one class of saturable binding sites with a dissociation constant of KD = 5.29 +/- 1.26 nM (mean +/- SE) and maximal binding Bmax = 357 +/- 113 fmol/10(6) cells. Competition studies showed that IC50 for Ro5-4864 was 15.7 nM, for PK11195 was 10.9 nM, for diazepam was 191 nM, and for clonazepam was greater than 10(-5) M. Studies in RBL-1 cells also showed one class of saturable binding sites with KD = 1.11 +/- 0.72 nM and Bmax = 177 +/- 69 fmol/10(6) cells; specificity was similar to peritoneal mast cells. Incubation of peritoneal mast cells with Ro5-4864 (10(-10)-10(-5) M) for 60 min, or with 10(-6) M for 0-60 min, showed no change in histamine or serotonin release either in unstimulated mast cells or those stimulated by compound 48/80 or substance P. Experiments with the fluorescent dye Quin-2 showed no change in free intracellular calcium levels in peritoneal mast cells after incubation with Ro5-4864 ranging from 10(-10) to 10(-6) M.

Animals↗
PubMed1988

Peripheral type benzodiazepine receptor and airway smooth muscle relaxation.

It has been postulated that a benzodiazepine receptor with a micromolar affinity may be associated with Ca++ channels in peripheral organs. We examined the actions of Ro5-4684 (parachlorodiazepam) and midazolam on guinea pig tracheal smooth muscle contraction. Binding studies using [3H]Ro5-4684 indicate the presence of a "peripheral" type binding site with a Kd of approximately 4 nM and maximum binding of 1 pmol/mg of protein. Midazolam did not displace radioligand. In tension studies no activity was seen for Ro5-4684 or midazolam at concentrations below 1 microM. Higher concentrations relaxed the airway smooth muscle under basal tone, the effect was augmented significantly by epithelium removal. Similar results were obtained in tissues precontracted with methacholine or KCl. Midazolam (1 or 100 microM) significantly (P less than .05) attenuated the response to Ca++ in K+-depolarized tracheal strips, the effect was greater at low Ca++ concentrations. The compounds appear to function as Ca++ antagonists in airway smooth muscle but ar not typical as shown by their ability to reduce basal tone in airway smooth muscle.

Animals↗
PubMed1988

Chronic benzodiazepine administration. II. Discontinuation syndrome is associated with upregulation of gamma-aminobutyric acidA receptor complex binding and function.

A "withdrawal" or "rebound" syndrome may follow the abrupt discontinuation of chronic treatment with benzodiazepines. To assess the neurochemical basis of this syndrome, mice were treated with lorazepam, 2 mg/kg/day for 7 days, a course which produces tolerance and downregulation of receptor binding and function. Behavioral studies indicated that open-field activity in lorazepam-treated mice was similar to controls at 1 day postlorazepam. Activity was increased at 4 days postlorazepam, and had returned to control levels by day 7. Benzodiazepine receptor binding as measured by specific uptake of [3H]Ro15-1788 in several brain regions returned to control levels by day 2 postbrazepam but had increased above controls at day 4 postlorazepam. Binding returned to control levels by day 7. Similar results were observed using [3H]flunitrazepam in membrane preparations. A similar time course was also observed for the maximum binding of the chloride channel ligand t-[35S]butylbicyclophosphorothionate during this period. The IC50 value for muscimol inhibition of t-butylbicyclophosphorothionate binding was decreased at day 4, suggesting increased coupling between gamma-aminobutyric acid and the chloride channel. Muscimol-stimulated [36Cl-] up-take in cortical synaptoneurosomes was increased at day 4 postlorazepam. These results indicate that the benzodiazepine discontinuation syndrome is associated with upregulation of receptor binding and function at the gamma-aminobutyric acidA receptor complex.

Animals↗
PubMed1988

Chronic benzodiazepine administration. I. Tolerance is associated with benzodiazepine receptor downregulation and decreased gamma-aminobutyric acidA receptor function.

Tolerance occurs to a number of the pharmacodynamic effects of benzodiazepines. To assess pharmacokinetic and neurochemical aspects of tolerance, lorazepam (LRZ) was administered chronically to mice via implantable osmotic pumps and rotarod ataxia, plasma and brain LRZ concentrations, benzodiazepine receptor binding in vivo and in vitro, chloride channel binding and muscimol-stimulated chloride uptake were examined in various brain regions over a 14-day period. Behavioral tolerance, indicated by diminished rotarod ataxia, developed at all doses examined (1, 2, 4 and 10 mg/kg/day), with little change occurring before day 4. The greatest decrease in rotarod ataxia occurred between days 4 and 7. Plasma and brain LRZ concentrations were proportional to dose and were constant over time at each dose, indicating that tolerance was not pharmacokinetic. Benzodiazepine receptor binding as determined by the specific uptake of [3H]Ro15-1788 decreased in cortex, hypothalamus and hippocampus primarily between days 4 and 7, with an approximately 50% decrement in each region by day 7. Receptor binding and rotarod ataxia in cortex were highly correlated at each dose. Apparent affinity in vivo at the receptor was unchanged in cortex, indicating that altered ligand uptake was due to decreased receptor number. Similar results were observed in membrane preparations. There was a small, nonsignificant decrease in chloride channel binding at day 7 compared to day 1. Muscimol-stimulated chloride uptake into cortical synaptoneurosomes was decreased at day 7 compared to day 1. Thus, downregulation of benzodiazepine receptor binding and of gamma-aminobutyric acidA receptor function is closely associated with behavioral tolerance to benzodiazepines.

Animals↗
PubMed1988

Structural specificity of MIF-1 and Tyr-MIF-1 in augmenting GABA-stimulated benzodiazepine receptor binding.

The possibility of a linkage between endogenous peptides and the GABA-benzodiazepine system has been greatly strengthened by reports that the brain peptides MIF-1(Pro-Leu-Gly-NH2) and Tyr-MIF-1(Tyr-Pro-Leu-Gly-NH2) augment GABA-stimulated benzodiazepine binding. We evaluated the actions of fragments and analogs of these small peptides on GABA-stimulated benzodiazepine binding in mouse cortical brain membranes. Fragments of Tyr-MIF-1, several compounds related to MIF-1, or several analogs of Tyr-MIF-1 had no effect on binding. These results demonstrate the specificity of the effects of MIF-1 and Tyr-MIF-1 on GABA-stimulated benzodiazepine binding.

Animals↗
PubMed1987

Rapid increase in brain benzodiazepine receptor binding following defeat stress in mice.

Defeat stress in mice, a model of social stress, increases benzodiazepine receptor binding as measured by specific [3H]Ro15-1788 binding in vivo, but not by [3H]flunitrazepam binding in vitro. This increase occurs rapidly, by 20 min following exposure to stress, and resolves by 60 min. Increased benzodiazepine receptor binding is observed in the cerebral cortex, cerebellum and hypothalamus, and appears to be due to an increase in receptor number rather than apparent affinity. The stress-induced increase in central benzodiazepine receptors is decreased in a dose-dependent fashion by lorazepam, a benzodiazepine agonist, but not by the receptor antagonist Ro15-1788. The stress-induced increase in benzodiazepine receptors is also blocked by adrenalectomy and is restored by corticosterone replacement.

Adrenalectomy↗
PubMed1987

Actual versus reported benzodiazepine usage by medical outpatients.

Benzodiazepines are widely prescribed, and in 1979 almost 10% of the adult population was taking them. Prior studies of outpatient usage of benzodiazepines have relied on survey or prescription data, which may be confounded by noncompliance. To determine the actual use of benzodiazepines, plasma benzodiazepine concentrations were measured in 225 consecutive outpatients from a university cardiology outpatient service. Self reports indicated that the great majority of the patients (191) were taking at least one medicine, and 70 reported being on a psychotropic drug. Seventy-seven patients reported taking benzodiazepines, the majority being on bromazepam (20), diazepam (26) or oxazepam (19). In 25 of those 77 patients, the reported drug could not be detected in plasma. Conversely, in 10 of the 225 patients, benzodiazepines which were not reported were detected (diazepam or flurazepam). Of those taking benzodiazepines, many had a low concentration, suggesting intermittent rather than regular use. Thus, many patients for whom benzodiazepines are prescribed take them irregularly, and a small group uses them without reporting their prescription. These findings have implications for the clinical presentation of illness and for the possibility of drug interactions.

Adolescent↗
PubMed1987

Tyr-MIF-1 augments benzodiazepine receptor binding in vivo.

Behavioral and limited neurochemical evidence indicates possible links between the endogenous opiate and gamma-aminobutyric acid (GABA)-benzodiazepine receptor systems. A previous study using in vitro techniques indicated that MIF-1 (Pro-Leu-Gly-NH2) and Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2), peptides with anti-opiate activity, enhanced GABA-stimulated benzodiazepine binding. To assess the activity of these peptides under in vivo conditions, we evaluated their effects on benzodiazepine receptor binding as determined by specific uptake of the benzodiazepine antagonist [3H]-Ro15-1788. Tyr-MIF-1, at a dose of 1 mg/kg IP, significantly augmented benzodiazepine binding in cortex and hippocampus but not in cerebellum, hypothalamus, or pons-medulla. Increases in binding were due in large part to increased apparent affinity at the receptor. At none of the doses of MIF-1 (0.1, 1 and 10 mg/kg) or at the highest (10 mg/kg) and lowest (0.1 mg/kg) doses of Tyr-MIF-1 was there any significant alteration in benzodiazepine binding in any region evaluated. These results indicate that peptide-benzodiazepine receptor interactions may also occur in vivo.

Animals↗
PubMed1987

Effects of Tyr-MIF-1 and MIF-1 at the GABAA receptor chloride channel site.

The peptides Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2) and, to a lesser extent, MIF-1 (Pro-Leu-Gly-NH2) recently have been found to augment the effects of gamma-aminobutyric acid (GABA) on benzodiazepine receptor binding at the GABAA receptor complex. To assess their interaction with the chloride channel binding site on the GABAA receptor, we evaluated the effects of these two peptides on [35S]-t-butylbicyclophosphorothionate (TBPS) binding in mouse brain membranes. In cortex, neither peptide altered [35S]-TBPS binding over a broad dose range, but Tyr-MIF-1 significantly augmented displacement of radioligand binding by the GABA analog muscimol at peptide concentrations of 10(-10) to 10(-7) M; MIF-1 had little effect on muscimol displacement of [35S]-TBPS binding. In cerebellum and brainstem, neither peptide was active in altering muscimol displacement of binding. Thus, Tyr-MIF-1 augments the displacement of [35S]-TBPS binding by the GABA analog muscimol in mouse brain cortical membranes, indicating that this peptide enhances the effects of GABA at the chloride channel as well as at the benzodiazepine receptor.

Animals↗