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Biomedical subjects

R A Harris

Publications and source records attributed to R A Harris.

At least 181 records · Page 10Linked to original sources
PubMed1996

Modulation of GABAA receptor function by G protein-coupled 5-HT2C receptors.

Two classical neurotransmitters, 5-hydroxytryptamine (5-HT) and GABA, coexist in neurons of the medulla oblongata, and activation of 5-HT receptors modulates GABAA receptor function in neurons of the ventral tegmental area, substantia nigra and cerebellum. We now report that activation of 5-HT2C receptors produces a long-lasting (20-90 min) inhibition of GABAA receptors in Xenopus oocytes coexpressing both types of receptors 5-HT2C receptors caused a approximately 60% decrease in the GABAA receptor Emax without affecting the EC50 or Hill coefficient. Intracellular microinjection of 500 microM BAPTA blocked, whereas microinjection of inositol 1,4,5-triphosphate mimicked the inhibitory action of 5-HT2C receptors. The inhibition was independent of the GABAA receptors subunit composition; receptors containing alpha 2 beta 1, alpha 1 beta 1 gamma 2L, and alpha 2 beta 1 gamma 2S were inhibited to the same extent by 5-HT2C receptor activation. Moreover, GABAA receptors composed of wild-type alpha 2 plus mutant beta 1(S409A) subunits were inhibited to the same extent as wild-type receptors. The nonspecific protein kinase inhibitor, staurosporine, and the inhibitor of serine/threonine protein phosphatases, calyculin A, did not block the inhibitory effects of 5-HT2C receptors. The results with these inhibitors, taken together with those obtained with GABAA receptors with different subunit compositions, suggest that protein kinases or serine/threonine phosphatases are not involved in this GABAA receptor modulatory process. Thus, we propose that 5-HT2C receptors inhibit GABAA receptors by a Ca(2+)-dependent, but phosphorylation independent, mechanism and that 5-HT and GABA may act as cotransmitters to regulate neuronal activity. Furthermore, disruption of the cross-talk between these receptors may play a role in the anti-anxiety actions of 5-HT2 receptor antagonists.

Animals↗
PubMed1996

The cytoskeleton and neurotransmitter receptors.

The neuronal cytoskeleton consists of microtubules and microfilaments that can interact with membrane proteins including neurotransmitter receptors and ion channels. Ligand-gated ion channels, such as nicotinic acetylcholine receptors, glycine receptors, glutamate receptors and gamma-aminobutryic acidA (GABAA) receptors, are known to cluster in plasma membranes. Studies suggest that postsynaptic ligand-gated channels form clusters that are anchored in the plasma membrane by interacting with cytoskeletal components and these clusters may serve to optimize delivery of neurotransmitters to the channels. Other findings indicate that the interaction of clustered ligand-gated ion channels with cytoskeletal components may also play a role in channel function. For example, studies suggest that the interaction of microtubules with GABAA receptors regualtes GABA binding affinity. Regulation of neurotransmitter function may be significant in the study of neuropathological processes, such as Alzheimer's disease, neurotrauma, and experimental epilepsy, in which the cytoskeleton is vulnerable to disruption.

Animals↗
PubMed1996

Cytokine production in hearts of Trypanosoma cruzi-infected CBA mice: Do cytokine patterns in chronic stage reflect the establishment of myocardial pathology?

The authors analysed cytokine production in hearts of Trypanosoma cruzi-infected CBA-J mice by in situ immunocytochemical staining. Cellular infiltrates were recorded in hearts from both acute and chronic stages, but were not apparent in control hearts. In the acute heart, CD8 cells predominated, with associated production of IL-4, IL-6 and TNF-alpha. Cytokine production was characterized by IL-4, IL-5, IL-6 and TNF-alpha in the chronic heart, and numbers of CD4 and CD8 cells were more equal. At this stage, calcified infarctions and associated fibrosis were apparent, mimicking chronic human Chagas' heart pathology. The authors consider the CBA mouse an appropriate model of chronic T. cruzi infection, and suggest that local cytokine production reflects establishment of heart pathology.

Acute Disease↗
PubMed1996

Microtubule depolymerization inhibits ethanol-induced enhancement of GABAA responses in stably transfected cells.

We studied whether microtubule organization is important for actions of ethanol on GABAA ergic responses by testing the effects of microtubule depolymerization on ethanol enhancement of GABA action in mouse L(tk-) cells stably transfected with GABAA receptor alpha 1 beta 1 gamma 2L subunits. The microtubule-disrupting agents colchicine, taxol, and vinblastine completely blocked ethanol-induced enhancement of muscimol-stimulated chloride uptake. beta-Lumicolchicine, a colchicine analogue that does not disrupt microtubules, had no effect on ethanol action. Colchicin did not alter the potentiating actions of flunitrazeparn or pentobarbital on muscimol-stimulated chloride uptakes. Thus, colchicine specifically inhibited the potentiating action of ethanol. From these findings, we conclude that intact microtubules are required for ethanol-induced enhancement of GABAA responses and suggest that a mechanism involving microtubules produces posttranslational modifications that are necessary for ethanol sensitivity in this cell system.

Animals↗
PubMed1996

Transfected cells for study of alcohol actions on GABA(A) receptors.

The ability to transfect and express foreign genes in cultured cells and Xenopus oocytes has contributed immensely to our knowledge of drug-receptor interactions and signal transduction following receptor activation. These techniques are now being used to study acute and chronic effects of ethanol on neurotransmitter receptors and cellular signaling. This article reviews the use of transfected cells to study ethanol actions on the GABAA receptor.

Journal Article↗
PubMed1996

Hepatic cystic disease in an adult polycystic kidney disease transplant population.

BACKGROUND: A study of the incidence of polycystic liver in a transplant population and an assessment of the impact of this disease on the group was undertaken. Clinical presentation, investigation and treatment of hepatic polycystic disease are explored. METHODS: The study examined the morbidity incurred by polycystic liver disease in patients, patient and graft survival, incidence of hepatic cysts and presentation, investigation and management of morbid hepatic cystic disease. One hundred and eleven patients were studied. RESULTS: It was found that hepatic cysts occurred in the majority of patients that underwent transplants for renal failure because of polycystic kidney disease at this institution. Symptomatic hepatic cystic disease was found to be primarily responsible for three deaths and affected between 15 and 20% of the group. Gall bladder disease was found to be associated with highly symptomatic hepatic cystic disease. CONCLUSION: Hepatic cystic disease is common in patients that have had kidney transplants because of polycystic kidney disease. Morbidity is likely to increase as patients survive for longer periods. Cholecystectomy should be considered in patients with significant hepatic cystic disease.

Adult↗
PubMed1996

Cellulitis and the occult diabetic.

BACKGROUND: Occult diabetes may be an important factor in the development of cellulitis and cellulitis may act as a precipitant to the diagnosis of diabetes mellitus. The present study defines the prevalence of diabetes and glucose tolerance impairment in a group presenting with cellulitis to a teaching hospital. A description of the demographic and pathological presentation of the group is undertaken. METHODS: Five hundred consecutive admissions for cellulitis to a Sydney teaching hospital were analysed. The cases presented between 1985 and 1994. Precipitating factors, length of stay, site of infection, white cell count, degree of fever, blood sugar estimation, history of diabetes mellitus and microbiological diagnosis were recorded. RESULTS: Forty-nine patients had a prior diagnosis of diabetes mellitus. Twenty-one percent of patients (56/265) were noted to have abnormal glucose tolerance on routine testing. Thirty-seven per cent of the 1994 cohort (14/38) demonstrated abnormal glucose tolerance. The most common precipitant was trauma (137/500). Mean length of stay was 8.7 days. Microbiological diagnosis was made in 32% of cases. CONCLUSIONS: Cellulitis requiring admission to hospital is a significant problem in terms of cost and bed occupation. A presentation with cellulitis may be a clinical indicator of impaired glucose tolerance. All patients presenting with cellulitis should have a fasting blood sugar level determined as part of routine workup.

Adult↗
PubMed1996

Actions of long chain alcohols on GABAA and glutamate receptors: relation to in vivo effects.

1. The effects of n-alcohols on GABAA and glutamate receptor systems were examined, and in vitro effectiveness was compared with in vivo effects in mice and tadpoles. We expressed GABAA, NMDA, AMPA, or kainate receptors in Xenopus oocytes and examined the actions of n-alcohols on receptor function using two-electrode voltage clamp recording. 2. The function of GABAA receptors composed of alpha 1 beta 1 or alpha 1 beta 1 gamma 2L subunits was potentiated by all of the n-alcohols studied (butanol-dodecanol). 3. In contrast to GABAA receptors, glutamate receptors expressed from mouse cortical mRNA or from cRNAs encoding AMPA (GluR3)- or kainate (GluR6)-selective subunits were much less sensitive to longer chain alcohols. In general, octanol and decanol were either without effect or high concentrations were required to produce inhibition. 4. In contrast to the lack of behavioural effects by long chain alcohols reported previously, decanol produced loss of righting reflex in short- and long-sleep mice, indicating that the in vivo effects of decanol may be due in part to actions at GABAA receptors. Furthermore, butanol, hexanol, octanol, and decanol produce similar potentiation of GABAA receptor function at concentrations required to cause loss of righting reflex in tadpoles, an in vivo model where alcohol distribution is not a compromising factor. 5. Thus, the in vivo effects of long chain alcohols are not likely to be due to their actions on NMDA, AMPA, or kainate receptors, but may be due instead to potentiation of GABAA receptor function.

Alcohols↗
PubMed1996

Enhancement of homomeric glycine receptor function by long-chain alcohols and anaesthetics.

1. The effects of n-alcohols (ethanol to dodecanol) and anaesthetics on strychnine-sensitive glycine receptors were studied in Xenopus oocytes expressing homomeric alpha 1 or alpha 2 glycine receptor subunits, with the two electrode voltage-clamp recording technique. 2. The glycine-induced chloride conductance of homomeric alpha glycine receptors was potentiated by all the alcohols tested when an EC2 concentration of glycine was used. Homomeric alpha 1 and alpha 2 receptors were potentiated similarly by the n-alcohols, except that low concentrations of ethanol produced greater potentiation with alpha 1, as previously reported. 3. Increasing the n-alcohol carbon number has been shown to increase the potency of the alcohols up to decanol at concentrations corresponding to EC50s for producing loss of righting reflex in tadpoles. However, dodecanol was no more potent than decanol, and only modest potentiation (30-60%) was obtained with dodecanol, in contrast to marked (150-200%) potentiation with the other alcohols. Thus, a "cut-off' occurred at about dodecanol. 4. Propofol, alphaxalone, pentobarbitone, halothane and enflurane, reversibly potentiated the function of homomeric alpha 1 glycine receptors at concentrations which represent approximately twice the EC50 for production of anaesthesia in mammals, but ketamine and etomidate were ineffective. 5. Two novel cyclobutane compounds were tested; the anaesthetic compound (1-chloro-1,2,2-trifluorocyclobutane) from 0.5 to 5 mM potentiated the action of glycine in a concentration-dependent manner; however, the non-anaesthetic analogue (1,2-dichloro-hexfluorocyclobutane) had no effect on glycine receptor function at concentrations (25 to 80 microM) predicted to be anaesthetic, based on the lipid solubility of this compound. 6. These results suggest that the alpha subunits of strychnine-sensitive glycine receptors contain sites of action for n-alcohols, propofol, alphaxalone, pentobarbitone and volatile anaesthetics, but not for ketamine and etomidate. Potentiation of glycine receptor function may contribute to the anaesthetic action of n-alcohols and volatile agents.

Anesthetics↗
PubMed1996

Regulation of GABAA receptor structure and function by chronic drug treatments in vivo and with stably transfected cells.

In this article, we review the use of stably transfected cells to study the regulation of receptor structure and function by chronic drug treatments and compare results from these cells to results obtained from other systems, including neuronal cultures and intact animals. We focus on the gamma-aminobutyric acid type A (GABAA) receptor complex. Sedative/hypnotic drugs such as benzodiazepines, barbiturates and alcohol that potentiate GABAA receptor function produce tolerance and dependence. Chronic treatment of GABAA receptor preparations from brain and neuronal cultures with GABAA agonists, as well as these other three classes of drugs, results in regulation of several properties of the receptor. Drug treatments may regulate levels of binding sites, allosteric binding interactions, receptor function, levels of receptor subunit mRNA and levels of receptor subunit protein. Some or all of these effects may comprise the molecular mechanisms of tolerance to these GABAA-modulatory drugs. The use of cells stably transfected with neurotransmitter receptors provides a homogeneous population that can be cultured under controlled conditions. As most preparations contain mixed populations of GABAA receptor subunits, stably transfected cells offer the advantage of the expression of receptors with a defined subunit composition. We conclude that chronic drug treatments regulate allosteric coupling and function of GABAA receptors in stably transfected cells. This regulation does not appear to be due to decreases in the expression of alpha 1- or beta 1-receptor subunits or to expression of subunits other than alpha 1, beta 1, gamma 2L. Therefore, it is unlikely to be due to changes in receptor subunit composition and probably represents post-translational changes. The rapid regulation of allosteric coupling and function by drug treatment of the stably transfected cells should provide insights to the mechanisms of coupling between GABAA and benzodiazepine receptors as well as tolerance and dependence of benzodiazepines and ethanol.

Animals↗
PubMed1996

Inhibition of rho1 receptor GABAergic currents by alcohols and volatile anesthetics.

We studied the effects of alcohols and anesthetics on homomeric gamma-aminobutyric acid (GABA) receptors formed of rho1 subunits expressed in Xenopus laevis oocytes. This subunit shares considerable amino acid sequence homology with the GABA(A) receptor subunits. In contrast to our previous findings with a variety of GABA(A) receptors, ethanol (10-100 mM) significantly inhibited the current induced by a low concentration (400 nM) of GABA, in an apparently competitive manner. Butanol (2-40 mM), hexanol (0.5-4 mM), heptanol (0.3-1 mM), octanol (0.055-1 mM) and nonanol (45 and 113 microM) also inhibited these GABAergic currents. Although efficacious positive modulators of GABA(A) receptor function, the volatile anesthetics enflurane, halothane and isoflurane inhibited the function of rho1 receptors. All of these drug effects were fully reversed by a 6-min washout period. When higher concentrations of GABA were used (5 microM) producing approximately 80-90% of a maximal response) neither the alcohols nor enflurane had any effect. In agreement with previous reports, pentobarbital (50 and 200 microM) was ineffective at any GABA concentration tested. Alphaxalone and propofol were also without effect. Furthermore, none of these compounds produced any significant effects when applied to oocytes in the absence of exogenously added GABA. The opposite effects of alcohols and anesthetics on rho1 and GABA(A) receptors, despite their significant amino acid sequence homology, may help in the identification of the particular amino acids responsible for the actions of these compounds on these receptors.

Alcohols↗
PubMed1996

A single amino acid determines differences in ethanol actions on strychnine-sensitive glycine receptors.

Effects of ethanol on strychnine-sensitive glycine receptors were studied in Xenopus laevis oocytes expressing alpha 1 wild-type, alpha 2, or mutant alpha 1(A52S) homomeric glycine receptors. This alpha 1(A52S) mutant, in which a serine residue substitutes for alanine at amino acid 52, is responsible for the spasmodic phenotype in mice and alters the ability of glycine to activate the receptor. Pharmacologically relevant concentrations of ethanol (10-200 mM) reversibly potentiated the glycine receptor function in all receptors. Ethanol potentiation depended on the glycine concentration used, with decreased potentiation observed at higher glycine concentrations. Homomeric alpha 1 glycine receptors were more sensitive to the effects of ethanol than were alpha 2 or the mutant alpha 1(A52S) receptors. No differences were found in ethanol sensitivity between alpha 2 and the mutant alpha 1(A52S) receptors. The alpha 2 subunit has a threonine residue, a conservative substitution for serine, at amino acid 52. The general anesthetic propofol was also tested in homomeric alpha 1, alpha 2, or the mutant alpha 1(A52S) receptors. Propofol, at unaesthetic concentrations (1-5 microM), reversibly potentiated the glycine receptor function in a concentration-dependent manner and to an equal extent in the three subunits tested. These data suggest that the mutation of an alanine to serine at amino acid 52 of the alpha subunit is responsible for the difference in ethanol sensitivity seen in homomeric receptors composed of alpha 1 and alpha 2 subunits.

Animals↗
PubMed1996

Anesthetics produce subunit-selective actions on glutamate receptors.

We assessed the involvement of specific glutamate receptors in the action of anesthetics. In addition to the clinical anesthetics enflurane, isoflurane and halothane, we tested novel halogenated compounds, which are anesthetic or nonanesthetic in vivo, on glutamate receptor (GluR) subunits. these volatile compounds as well as pentobarbital and phenobarbital were tested on kainate-induced currents in Xenopus oocytes expressing GluR1, GluR3, GluR2+3 or GluR6 subunits. The anesthetic 1-chloro-1,2,2-triflurocyclobutane (F3) weakly inhibited kainate responses in oocytes expressing GluR3 receptors but not oocytes expressing GluR1 or GluR2+3 receptors. Surprisingly, F3 potentiated kainate responses in oocytes expressing the kainate-selective receptor GluR6. The nonanesthetics 2,3-chlorooctafluorobutane (F8) and 1,2-dichlorohexafluorocyclobutane (F6) did not affect GluR3 or GluR6 kainate responses. Isoflurane weakly inhibited although enflurane and halothane modestly inhibited kainate responses in oocytes expressing GluR1, GluR3 or GluR2+3 receptors. As with F3, isoflurane, enflurane and halothane potentiated kainate-induced currents of GluR6 receptors. The respective inhibitory and potentiating effects of halothane on GluR3 and GluR6 receptors were enhanced by increasing duration of halothane exposure. In contrast to the opposing action of volatile anesthetics, pentobarbital and phenobarbital inhibited GluR3 and GluR6 kainate responses and had a much greater effect on GluR6 receptors. These results provide novel evidence that anesthetics produce selective actions on glutamate receptors, suggesting that subunit composition may determine the role of glutamate receptors in anesthesia.

Anesthetics↗
PubMed1995

Roles of amino acid residues surrounding phosphorylation site 1 of branched-chain alpha-ketoacid dehydrogenase (BCKDH) in catalysis and phosphorylation site recognition by BCKDH kinase.

Branched-chain alpha-ketoacid dehydrogenase is regulated by reversible phosphorylation of serine 293 (site 1) on the E1 alpha subunit. Alanine-scanning mutagenesis was used to examine the roles of residues surrounding serine 293 in catalysis by the dehydrogenase and in substrate recognition by branched-chain alpha-ketoacid dehydrogenase kinase. Alanine substitution of serine 293 resulted in a 10-fold increased Km for alpha-ketoisovalerate, a less increased (2.8-fold) Km for alpha-ketoisocaproate, but no change in Vmax or the Km for thiamine pyrophosphate. Alanine substitutions of arginine 288, histidine 292, and aspartate 296, residues highly conserved among alpha-ketoacid dehydrogenases, resulted in inactive enzymes. Each of the inactive E1 mutants bound to the E2 core subunit with equal affinity as wild-type E1, and each produced circular dichroism spectra identical to that of wild-type E1. Two mutations, H292A and S293E, abolished the ability of E1 apoenzyme to reconstitute with thiamine pyrophosphate. Each alanine-substituted E1 was phosphorylated at site 1 by branched-chain alpha-ketoacid dehydrogenase kinase with similar rates, with the exception of the R288A mutant, which displayed no detectable phosphorylation. Thiamine pyrophosphate inhibited the phosphorylation of all mutant enzymes with the exception of H292A, the mutant E1 that did not bind thiamine pyrophosphate.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1995

The role of coronary angiography and coronary revascularization before noncardiac vascular surgery.

OBJECTIVE: To determine whether preoperative coronary angiography and revascularization improve short-term outcomes in patients undergoing noncardiac vascular surgery. DESIGN: Decision analysis. PATIENTS: Patients undergoing elective vascular surgery who had either no angina or mild angina and a positive dipyridamole-thallium scan result. INTERVENTIONS: Three strategies were compared. The first strategy was to proceed directly to vascular surgery. The second was to perform coronary angiography, followed by selective coronary revascularization, before proceeding to vascular surgery and to cancel vascular surgery in patients with severe inoperable coronary artery disease (CAD). The third was to perform coronary angiography, followed by selective coronary revascularization, before proceeding to vascular surgery and to perform vascular surgery in patients with inoperable CAD. MAIN OUTCOME MEASURES: Mortality, nonfatal myocardial infarction, stroke, uncorrected vascular disease, and cost. All outcomes were assessed within 3 months. RESULTS: Proceeding directly to vascular surgery led to lower morbidity and cost in the base case analysis. The coronary angiography strategy led to higher mortality if vascular surgery would proceed in patients with inoperable CAD, but led to slightly lower mortality if vascular surgery were canceled in patients with inoperable CAD. The coronary angiography strategy also led to lower mortality when vascular surgery was particularly risky. CONCLUSIONS: Decision analysis indicates vascular surgery without preoperative coronary angiography generally leads to better outcomes. Preoperative coronary angiography should be reserved for patients whose estimated mortality from vascular surgery is substantially higher than average.

Coronary Angiography↗
PubMed1995

Screening surgeons for HIV infection. A cost-effectiveness analysis.

UNLABELLED: OBJECTIVE. To determine the cost-effectiveness of a policy to screen surgeons for human immunodeficiency virus (HIV) infection to prevent transmission of HIV to patients having invasive procedures. DESIGN: Cost-effectiveness analysis. RESULTS: A one-time national screening program would identify approximately 137 surgeons with HIV infection (range, 28 to 423 surgeons) and would prevent approximately 4.3 infections (range, 1.9 to 21.3 infections) in patients treated by infected surgeons and 0.9 infections (range, 0 to 12.9 infections) in sexual partners of infected surgeons at a direct cost of $8.1 million and an induced cost of approximately $44 million. It would result in expenditures of $458,000 per year of life saved (range, $147,000 to $687,000 per year of life saved), whereas an annual screening program would result in expenditures of approximately $1.1 million per year of life saved (range, $338,000 to $1,886,000 per year of life saved). If the prevalence of HIV infection in surgeons is estimated to be three times our base-case estimate (an increase from 0.1% to 0.3%), annual screening would result in expenditures of approximately $741,000 per year of life saved. If the probability of seroconversion after a patient is exposed to a contaminated instrument is increased to 5.0% from our base-case estimate of 0.29%, an annual screening program would still cost more than $228,000 per year of life saved. CONCLUSION: Screening surgeons for HIV to prevent transmission of HIV to patients having invasive procedures requires expenditures per year of life saved that are considerably in excess of those of most accepted health interventions. Surveillance studies of patients treated by surgeons infected with HIV should be continued to confirm that transmission of HIV to patients having invasive procedures is rare.

Cost-Benefit Analysis↗