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Biomedical subjects

P Skolnick

Publications and source records attributed to P Skolnick.

At least 181 records · Page 10Linked to original sources
PubMed1989

1-Aminocyclopropane carboxylic acid: a potent and selective ligand for the glycine modulatory site of the N-methyl-D-aspartate receptor complex.

1-Aminocyclopropane carboxylic acid (ACPC) competitively inhibited (IC50, 38 +/- 7 nM) [3H]glycine binding to rat forebrain membranes but did not affect [3H]strychnine binding to rat brainstem/spinal cord membranes. Like glycine, ACPC enhanced 3H-labelled (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate ([3H]MK-801) binding to N-methyl-D-aspartate receptor-coupled cation channels (EC50, 135 +/- 76 nM and 206 +/- 78 nM for ACPC and glycine, respectively) but was approximately 40% less efficacious in this regard. The maximum increase in [3H]MK-801 binding produced by a combination of ACPC and glycine was not different from that elicited by glycine, but both compounds potentiated glutamate-stimulated [3H]MK-801 binding. These findings indicate that ACPC is a potent and selective ligand at the glycine modulatory site associated with the N-methyl-D-aspartate receptor complex.

Amino Acids↗
PubMed1989

GABAA receptor complex in an experimental model of hepatic encephalopathy: evidence for elevated levels of an endogenous benzodiazepine receptor ligand.

The involvement of the gamma-aminobutyric acidA (GABAA) receptor complex in the pathogenesis of hepatic encephalopathy was examined in thioacetamide-treated rats with fulminant hepatic failure. Partially purified extracts from encephalopathic rat brain were approximately three times more potent in inhibiting [3H]Ro 15-1788 binding to benzodiazepine receptors than identically prepared extracts from control rats. High levels of inhibitory activity were also found in extracts of plasma, heart, and liver from thioacetamide-treated rats. The inhibition of [3H]Ro 15-1788 binding by brain extracts appeared to be competitive and reversible and was unaffected by treatment with either proteolytic enzymes or boiling. Further, GABA significantly enhanced the potency of these extracts in inhibiting [3H]flunitrazepam binding. In contrast, no differences were found in radioligand binding to the constituent recognition sites of the GABAA receptor complex in well-washed brain membranes prepared from control and encephalopathic animals. These findings suggest that the recognition-site qualities of the constituent proteins of the GABAA receptor complex are unchanged in an experimental model of hepatic encephalopathy. However, significant elevations in the level of a substance or substances with neurochemical properties characteristic of a benzodiazepine receptor agonist may contribute to the electrophysiological and behavioral manifestations of hepatic encephalopathy.

Animals↗
PubMed1989

Characterization of the effects of AHN 086, an irreversible ligand of "peripheral" benzodiazepine receptors, on contraction in guinea-pig atrial and ileal longitudinal smooth muscle.

The effects of AHN 086 and its reversibly acting structural analogue Ro 5-4864 were studied in the spontaneously beating guinea-pig atria and field-stimulated guinea-pig ileal longitudinal smooth muscle in the presence and absence of dihydropyridine calcium channel modulators. The treatment of guinea-pig atria with AHN 086 followed by extensive washing did not alter contraction. However, AHN 086 (0.5 microM) potentiated (88%) the positive inotropic responses by BAY K 8644, an effect that was not reversed by extensive washing of the tissue. Higher concentrations of AHN 086 (greater than 2 microM) irreversibly inhibited the intropic, but not the chronotropic responses to BAY K 8644, nifedipine, and isoproterenol. Ro 5-4864 (10 microM) produced a reversible enhancement of the inotropic responses and block of the chronotropic responses to BAY K 8644. In guinea-pig ileal longitudinal smooth muscle, both AHN 086 and Ro 5-4864 reversibly inhibited field-stimulated contractions. Neither Ro 5-4864 nor AHN 086 affected the ability of nifedipine to inhibit field-stimulated contractions of ileal longitudinal smooth muscle. Treatment of intact atrial with 5 microM AHN 086 followed by extensive washing resulted in a significant inhibition (30-50%) of [3H]Ro 5-4864 binding to peripheral benzodiazepine receptors and of [3H]nitrendipine binding to voltage-operated calcium channels, but did not affect [3H]dihydroalprenolol binding to beta-adrenergic receptors on atrial membranes. The same treatment applied to intact ileal longitudinal smooth muscle affected neither [3H] (-)-quinuclidinyl benzilate binding to muscarine receptors nor [3H]nitrendipine binding, but did result in a significant inhibition (30-50%) of [3H]Ro 5-4864 binding to ileal longitudinal smooth muscle membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1989

Electrophysiological actions of Ro5-4864 on cerebellar Purkinje neurons: evidence for "peripheral" benzodiazepine receptor-mediated depression.

The effects of Ro 5-4864 (4'-chlorodiazepam; the archetypic peripheral benzodiazepine receptor ligand) were examined on the electrophysiological responses of rat cerebellar Purkinje neurons maintained in vitro. Ro 5-4864 produced a biphasic response (consisting of an increase followed by a decrease in spontaneous firing) in approximately 50% of the neurons studied. The remaining neurons responded to Ro 5-4864 application with decreased spontaneous activity. The EC50 and IC50 values for the excitatory and depressant responses to Ro 5-4864 were 490 and 450 nM, respectively. Preincubation with PK 11195 [N-methyl-N-(methyl-1-propyl)chloro-2-phenyl-1-isoquinoline-3-carboxamid e; a peripheral benzodiazepine receptor antagonist] reduced the potency of Ro 5-4864 to inhibit cell firing in a competitive fashion, but did not alter Ro 5-4864-elicited excitations. A similar effect was observed with the monocarboxylic acid anion transport inhibitor UK 5099 [alpha-cyano-beta-(1-phenylindol-3-yl)acrylate]. In contrast, the increases in cell firing elicited by t-butylbicyclophosphorothionate (a GABA-gated chloride channel blocker) were enhanced 6-fold by coperfusion with both Ro 5-4864 and PK 11195. These findings suggest that the Ro 5-4864 induced depressions of Purkinje neuron spontaneous activity is mediated by peripheral benzodiazepine receptors, whereas the excitation may result from modulation of the gamma-aminobutyric acid-gated chloride ionophore.

Animals↗
PubMed1989

meta- and para-isothiocyanato-t-butylbicycloorthobenzoate: irreversible ligands of the gamma-aminobutyric acid-regulated chloride ionophore.

The meta- and para-isothiocyanato derivatives of t-butylbicycloorthobenzoate (TBOB) were synthesized by catalytic reduction of the corresponding nitro compounds, followed by treatment with thiophosgene. p-NCS-TBOB (2) inhibited the binding of both [3H]TBOB and [35S]t-butylbicyclophosphorothionate (TBPS) with potencies (IC50 of 61 and 23 nM, respectively) similar to the parent compound. In contrast, the meta derivative (m-NCS-TBOB, 1) was more than 1 order of magnitude less potent (IC50 of 1588 and 149 nM, respectively). The IC50 values for both 1 and 2 were strongly dependent on the tissue concentration, in a manner characteristic of irreversible inhibitors. Moreover, preincubation of tissue with these compounds, followed by extensive washing, resulted in a concentration-dependent reduction in the number of [35S]TBPS binding sites and in the apparent affinity of this radioligand. Similar effects were not observed in tissues treated in identical fashion with either TBOB or picrotoxin. Preincubation with p-NCS-TBOB at concentrations that significantly inhibit [35S]TBPS or [3H]TBOB binding did not affect radioligand binding to either benzodiazepine or gamma-aminobutyric acid receptors. These findings suggest that m- and p-NCS-TBOB bind irreversibly to sites labeled by cage convulsants such as TBOB and TBPS, which are on or near GABA-gated chloride channels. p-NCS-TBOB should prove useful in determining the molecular characteristics of the benzodiazepine receptor-coupled GABA-gated chloride ionophore.

Animals↗
PubMed1989

Ultraviolet irradiation selectively disrupts the gamma-aminobutyric acid/benzodiazepine receptor-linked chloride ionophore.

The ability of UV light to affect radioligand binding and 36Cl-uptake at the gamma-aminobutyric acidA (GABAA) receptor-chloride channel complex was examined. Exposure to 302 nm UV light produced a rapid (t1/2 = 4 min) reduction in [35S]t-butylbicyclo-phosphorothionate binding (assayed in the presence of 200 mM chloride) to sites associated with the GABAA receptor-coupled chloride ionophore. Saturation analysis revealed that this effect could be attributed entirely to a decrease in the maximum number of binding sites. Exposure to UV irradiation at lower (254 nm) and higher (366 nm) wavelengths also inhibited [35S]t-butylbicy-clophosphorothionate binding, but the respective rates of inactivation were 8- and 27-fold slower, compared with 302 nm. Other anion-dependent interactions at the GABAA receptor complex were disrupted in a similar manner. In the absence of permeant anion, [3H]flunitrazepam binding to benzodiazepine receptors was unaffected by 302 nm UV irradiation, whereas chloride-enhanced [3H]flunitrazepam binding was inhibited markedly. In the presence of 250-500 mM chloride, [3H]methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate binding to benzodiazepine receptors was also inhibited after UV exposure. Basal 36Cl- uptake into synaptoneurosomes was nearly doubled after 15 min of exposure to 302 nm light, whereas pentobarbital- and muscimol-stimulated 36Cl- uptake were reduced significantly. UV irradiation at 302 nm appears to disrupt selectively the anion-dependent functional interactions at the GABAA receptor complex. The apparent wavelength specificity suggests that the gating structure (channel) may contain tryptophan and/or tyrosine residues vital to the regulation of anion movement through the ionophore portion of this supramolecular receptor-ion channel complex.

Animals↗
PubMed1989

Radiation inactivation reveals discrete cation binding sites that modulate dihydropyridine binding sites.

In low ionic strength buffer (5 mM Tris.HCl), the binding of [3H] nitrendipine to dihydropyridine calcium antagonist binding sites of mouse forebrain membranes is increased by both Na+ and Ca2+. Radiation inactivation was used to determine the target size of [3H]nitrendipine binding sites in 5 mM Tris.HCl buffer, in the presence and absence of these cations. After irradiation, [3H] nitrendipine binding in buffer with or without Na+ was diminished, due to a loss of binding sites and also to an increase in Kd. After accounting for radiation effects on the dissociation constant, the target size for the nitrendipine binding site in buffer was 160-170 kDa and was 170-180 kDa in the presence of sodium. In the presence of calcium ions, [3H]nitrendipine binding showed no radiation effects on Kd and yielded a target size of 150-170 kDa. These findings suggest, as in the case of opioid receptors, the presence of high molecular weight membrane components that modulate cation-induced alterations in radioligand binding to dihydropyridine binding sites.

Animals↗
PubMed1988

Differential regulation of mu-opiate receptors in heroin- and morphine-dependent rats.

Rats made dependent on heroin and morphine exhibit both qualitative and quantitative differences in the characteristics of radioligand binding to mu-opioid receptors in the central nervous system. In brain membranes prepared from control animals, [3H]dihydromorphine (DHM) binding was best described by a two-site model, while in morphine-dependent rats, [3H]DHM binding was best described by a single-site model. In contrast, [3H]DHM binding to membranes from heroin-dependent animals was best described by a two-site model, with an increased density of the high-affinity, and no change in the low-affinity population compared to controls. Furthermore, both the number of binding sites for [3H]DAGO (a ligand that selectively labels a population of high-affinity mu-opiate receptors) and the sensitivity of [3H]DHM to sodium ions was increased in heroin; but not in morphine-dependent rats. These studies demonstrate that opiate receptors are differentially regulated in heroin- and morphine-dependent animals. Such neurochemical changes in mu-opiate receptors may underlie differences in the behavioral and pharmacological profiles of heroin and morphine reported in man.

Animals↗
PubMed1988

Alcohols stimulate gamma-aminobutyric acid receptor-mediated chloride uptake in brain vesicles: correlation with intoxication potency.

A series of short-chain alcohols, including ethanol, were examined for their abilities to stimulate gamma-aminobutyric acid (GABA) receptor-mediated chloride uptake into isolated brain vesicles. All of the alcohols tested stimulated 36 chloride uptake, at concentrations that occur during acute intoxication, and their potencies in stimulating GABA receptor-mediated chloride uptake were highly correlated with both their intoxication potencies in rats (r = 0.96; P less than 0.0001) and their membrane/buffer partition coefficients (r = 0.91; P less than 0.0005). Thus, the activity of alcohols at the GABA receptor-coupled chloride ion channel appears to be related to their ability to enter hydrophobic regions of the neuronal membrane. These data suggest that the anxiolytic, sedative/hypnotic and intoxicating properties of ethanol may, in part, be mediated via an action at central GABA receptors.

Alcoholic Intoxication↗
PubMed1988

Purification and characterization of an endogenous protein modulator of radioligand binding to "peripheral-type" benzodiazepine receptors and dihydropyridine Ca2+-channel antagonist binding sites.

Acidified extracts of rat antral stomach chromatographed on octadecylsilane cartridges contained material that inhibited the binding of [3H]Ro 5-4864 (4'-chlorodiazepam) and [3H]nitrenidipine to "peripheral-type" benzodiazepine receptors and dihydropyridine Ca2+-channel antagonist binding sites respectively. This material reduced the apparent affinities of both radioligands without significantly affecting the maximum number of binding sites. In contrast, the binding of [3H]diazepam, [3H]Ro 15-1788 (ethyl-8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazo [1,5a][1,4] benzodiazepine-3-carboxylate), and [3H]3-carbomethoxy-beta-carboline to "brain-type" benzodiazepine receptors and [3H]dihydroalprenolol binding to beta-adrenergic receptors were unaffected by this material. Subsequent column chromatography on hydroxylapatite purified this material by greater than 2000-fold. This semi-purified substance was resolved by reverse phase HPLC as one u.v. adsorbing peak that inhibited both [3H]Ro 5-4864 and [3H]nitrendipine binding. The activity of this 16,000 dalton substance was destroyed completely by both heat treatment and pronase and partially reduced by trypsin. Furthermore, the inhibitory activity of this substance was enhanced by Ca2+ in a concentration-dependent fashion (0.1 to 10 mM). Comparison of TLC scans of 2-9,10[3H]dipalmitoyl-phosphatidylcholine incubated with either the HPLC purified material or authentic phospholipase A2(PLA2) (Naja naja) revealed that this substance has enzymatic properties indistinguishable from PLA2. These findings suggest that this endogenous protein may be a PLA2 isoenzyme which may modify both "peripheral-type" benzodiazepine receptors and dihydropyridine Ca2+-channel antagonist binding sites.

Animals↗
PubMed1988

Modulation of neurotransmitter metabolism by dihydropyridine calcium channel ligands in mouse brain.

The regional concentrations of dopamine, serotonin, dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindole acetic acid were measured in mouse brain following administration of the dihydropyridine calcium channel activator BAY K 8644, and antagonist, nifedipine. BAY K 8644 (1-8 mg/kg) produced dose- and time-dependent increases in dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindoleacetic acid concentrations in the caudate, without altering dopamine and serotonin levels. No changes in 5-hydroxyindoleacetic acid concentration were observed in the raphe nuclei, hypothalamus, hippocampus and frontal cortex. Nifedipine (4 mg/kg) blocked BAY K 8644- (2 mg/kg) elicited increases in dihydroxyphenylacetic acid in the caudate. Furthermore, a higher dose of nifedipine (8 mg/kg) decreased dihydroxyphenylacetic acid and homovanillic acid, but did not affect dopamine, serotonin or 5-hydroxyindoleacetic acid concentrations, while a lower dose of nifedipine (2 mg/kg) significantly increased serotonin, 5-hydroxyindoleacetic acid and homovanillic acid, but did not affect dopamine and dihydroxyphenylacetic acid concentrations. The findings that both BAY K 8644 and nifedipine affect neurotransmitter metabolism in vivo in a dose-, time- and brain region-dependent manner, suggest that high-affinity dihydropyridine calcium channel binding sites play an important role in regulating neurotransmitter turnover in the central nervous system.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1988

3-Ethoxy-beta-carboline: a high affinity benzodiazepine receptor ligand with partial inverse agonist properties.

3-Ethoxy-beta-carboline binds with high affinity to benzodiazepine receptors in the central nervous system (Ki approximately equal to 10.1, 15.3, and 25.3 nM in rat cerebellum, cerebral cortex, and hippocampus, respectively). This compound has pharmacological actions reminiscent of benzodiazepine receptor partial inverse agonists such as FG 7142 and 3-carboethoxy-beta-carboline. Thus, while not a convulsant, 3-ethoxy-beta-carboline potentiated the convulsant actions of pentylenetetrazole in mice. Furthermore, this compound reduced both the time spent and the total entries in the open arms of an elevated plus maze and also inhibited stress-induced ulcer formation, effects that are also observed with benzodiazepine receptor inverse agonists. These findings suggest that 3-ethoxy-beta-carboline is a partial inverse agonist at benzodiazepine receptors which may prove useful for in vivo studies since it has a higher affinity for benzodiazepine receptors and better solubility than the commonly used partial inverse agonist FG 7142. Furthermore, 3-ethoxy-beta-carboline appears to be less vulnerable to metabolic degradation than ester analogs with a similar pharmacological profile such as 3-carboethoxy-beta-carboline.

Animals↗
PubMed1988

Regulation of renal peripheral benzodiazepine receptors by anion transport inhibitors.

The in vitro and in vivo regulation of [3H]Ro 5-4864 binding to peripheral benzodiazepine receptors (PBR) by ion transport/exchange inhibitors was studied in the kidney. The potencies of 9-anthroic acid, furosemide, bumetanide, hydrochlorothiazide and SITS as inhibitors of [3H]Ro 5-4864 binding to renal membranes were consistent with their actions as anion transport inhibitors (Ki approximately equal to 30 - 130 microM). In contrast, spironolactone, amiloride, acetazolamide, and ouabain were less potent (Ki = 100-1000 microM). Administration of furosemide to rats for five days resulted in a profound diuresis (approximately equal to 350% increase in urine volume) accompanied by a significant increase in PBR density (43%) that was apparent by the fifth day of treatment. Administration of hydrochlorothiazide or Ro 5-4864 for five days also caused diuresis and increased renal PBR density. Both the diuresis and increased density of PBR produced by Ro 5-4864 were blocked by coadministration of PK 11195, which alone had no effect on either PBR density or urine volume. The equilibrium binding constants of [3H]Ro 5-4864 to cardiac membranes were unaffected by administration of any of these drugs. These findings suggest that renal PBR may be selectively modulated in vivo and in vitro by administration of ion transport/exchange inhibitors.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1988

The imidazobenzodiazepine Ro 15-4513 antagonizes methoxyflurane anesthesia.

Parenteral administration of the imidazobenzodiazepine Ro 15-4513 (a high affinity ligand of the benzodiazepine receptor with partial inverse agonist qualities) produced a dose dependent reduction in sleep time of mice exposed to the inhalation anesthetic, methoxyflurane. The reductions in methoxyflurane sleep time ranged from approximately 20% at 4 mg/kg to approximately 38% at 32 mg/kg of Ro 15-4513. Co-administration of the benzodiazepine receptor antagonist Ro 15-1788 (16 mg/kg) or the inverse agonists DMCM (5-20 mg/kg) and FG 7142 (22.5 mg/kg) blocks this effect which suggests that the reductions in methoxyflurane sleep time produced by Ro 15-4513 are mediated via occupation of benzodiazepine receptors. Moreover, neither DMCM (5-20 mg/kg) nor FG 7142 (22.5 mg/kg) reduced methoxyflurane sleep time which suggests this effect of Ro 15-4513 cannot be attributed solely to its partial inverse agonist properties. These observations support recent findings that inhalation anesthetics may produce their depressant effects via perturbation of the benzodiazepine/GABA receptor chloride channel complex, and suggest that Ro 15-4513 may serve as a prototype of agents capable of antagonizing the depressant effects of inhalation anesthetics such as methoxyflurane.

Anesthesia, General↗