DOCSLIB.ORG

The Effects of Midazolam and Sevoflurane on the GABAA Receptors with Alternatively Spliced Variants of the Γ2 Subunit

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
The Effects of Midazolam and Sevoflurane on the GABAA Receptors with Alternatively Spliced Variants of the Γ2 Subunit Korean J Anesthesiol 2011 February 60(2): 109-118 Experimental Research Article DOI: 10.4097/kjae.2011.60.2.109 The effects of midazolam and sevoflurane on the GABAA receptors with alternatively spliced variants of the γ2 subunit Woosik Eom1, Jung Min Lee2, Jeongmi Park2, Kyungho Choi3, Sung­Jun Jung4, and Hee­Soo Kim2 1Department of Anesthesiology and Pain Medicine, National Cancer Center, Goyang, 2Department of Anesthesiology and Pain Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, 3Specific Organs Cancer Branch, Research Institute, National Cancer Center, Goyang, 4Department of Physiology, Hanyang University College of Medicine, Seoul, Korea Background: Emergence agitation after sevoflurane anesthesia in children can be prevented by midazolam. Alternative splicing of the GABAA receptor changes with age. Therefore, we hypothesized that alternative splicing of the γ2 subunit affects the GABA current when applying sevoflurane and midazolam. Methods: We performed the whole­cell patch clamp technique on human embryonic kidney 293 cells that were transfected with α1β2γ2L or α1β2γ2S. The concentration­response relations were recorded for midazolam and sevoflurane, and the co­application responses were measured at concentrations of 1.5 nM, 15 nM and 300 nM of midazolam and 0.5%, 2.0% and 4.0% of sevoflurane. Each GABA current was compared with that produced by 5 μM of GABA. Results: The concentration­response relationships for midazolam and sevoflurane were dose­dependent without any differences between the α1β2γ2L and α1β2γ2S subtypes. 1.5 nM and 15 nM of midazolam did not significantly enhance the current after treatment with 0.5% sevoflurane for both subtypes. The current after treatment with 2.0% sevoflurane was enhanced by 1.5 nM midazolam for the α1β2γ2S subtype, but not for the α1β2γ2L subtype. In the case of 2.0% sevoflurane with 15 nM of midazolam, and 4.0% sevoflurane with 300 nM of midazolam, the GABA currents were significantly enhanced for both subtypes. Conclusions: These results show that the difference in the γ2 subunit cannot explain the emergence agitation after sevoflurane anesthesia in children in vitro. This suggests that co­application of sevoflurane and midazolam enhances the GABA current according to the alternative splicing of the γ2 subunit and the concentration of both drugs. (Korean J Anesthesiol 2011; 60: 109­118) Key Words: Agitation, Alternative splicing, GABAA receptor, Gamma 2 subunit, Midazolam, Sevoflurane. Received: August 5, 2010. Revised: August 19, 2010. Accepted: August 20, 2010. Corresponding author: Woosik Eom, M.D., Department of Anesthesiology and Pain Medicine, National Cancer Center, 323, Ilsan-ro, Goyang 410-769, Korea. Tel: 82-31-920-1703, Fax: 82-31-920-1463, E-mail: [email protected] CC This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http:// creativecommons.org/licenses/by-nc/3.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Copyright ⓒ the Korean Society of Anesthesiologists, 2011 www.ekja.org GABA receptor, midazolam and sevoflurane Vol. 60, No. 2, February 2011 Introduction receptor [10]. Sevoflurane is a volatile anesthetic that was developed The GABAA receptor mediates rapid inhibitory action by relatively recently, and it is now widely used and preferred for using GABA, which is a major inhibitory neurotransmitter of clinical anesthesia, and especially for pediatric anesthesia. the central nervous system. Activation of the GABAA receptor Sevoflurane has been reported to potentiate GABA­induced induces the opening of chloride channels and this increases currents at the GABAA receptor. Sevoflurane enables the rapid the inward chloride current. The hyperpolarization of the induction and quick recovery after inhalational anesthesia membrane of neurons then decreases the post­synaptic action when using it because sevoflurane possesses several favorable potential of the neuron [1]. properties, including low blood and tissue solubility, non­ As the GABAA receptor has specific binding sites for GABA, pungency and limited cardiorespiratory depression, which may barbiturate, benzodiazepines and the anesthetic steroids, which make sevoflurane desirable for use in infants and children [11]. induces sedation, hypnosis, amnesia or anesthesia through The quick recovery from sevoflurane is likely to be accompanied their binding, the GABAA receptor is considered to be a major by emergence agitation, which often occurs after administering target protein of anesthesia [2]. sevoflurane in children even when sufficient analgesia is The GABAA receptor is a complex of 5 subunits of 19 different provided [12,13]. subunits (α, β, γ, δ, ε, π, θ, ρ, etc.) in mammals. Multiple One of the trials to reduce the unpleasant effects during combinations from the different subunits make several subtypes emergence is to use midazolam as premedication. It has been of GABAA receptor. Most GABAA receptors consist of two α reported that premedication with oral midazolam is effective subunits, two β subunits and a γ subunit. The complexity of for decreasing the occurrence of emergence agitation without GABAA receptors is still under investigation and there exits the delaying the discharge from the post­anesthesia care unit and possibility of new combinations of subunits. Each combination so this is safe and suitable for outpatient surgery [14]. However, of different subunits has unique physiologic characteristics and the mechanism is still not clear, and it is not known whether the drug responses are different from each other [3]. sevoflurane and midazolam have the interaction at the level of The γ2 subunits of the GABAA receptor exist as a long type the GABAA receptor. (γ2L) and a short type (γ2S) by alternative splicing of RNA. The The mechanism of agitation was described in the model of γ2L subunits have another 8 amino acids (LLRMFSFK) in the Sachdev and Kruk as the result of decreased inhibitory signals intracellular loop and the phosphorylation by protein kinase C to the globus pallidus interna/substantia nigra pars or the (PKC) occurs in this region (Ser343) [4]. disinhibition of the thalamocortical neurons and brain stem It seems that the agents binding to the γ2 subunit may have neurons by the disturbance of neuronal circuits [15]. So, it is different effects according to the subtype. The total expression expected that the change of GABAA receptor activity may cause of the γ2 subunit is similar for all ages, but γ2L/γ2S increases or reduce emergence agitation. by the up­regulation of the γ2L subunit and γ2L becomes We postulated that alternative splicing of γ2 subunit is related dominant in mature synapses according to age [5,6]. with the occurrence of emergence agitation on the basis of the The distribution and pattern of GABAA receptors are various characteristics of midazolam, sevoflurane and the γ2 subunit; according to the region of the central nervous system and the sevoflurane binds to GABAA receptor, a benzodiazepine like type of neuron. This is also true for γ2L and γ2S [7,8]. The most midazolam prevents emergence agitation and binds to the α common pattern of GABAA receptor is the α1β2γ2 type, which is and γ subunits of the GABAA receptor, and alternative splicing 43% of all the GABAA receptors [9]. This implies that the diversity of the γ2 subunit is different according to age. Therefore, of the subtype variants and the distribution of GABAA receptors we performed the whole­cell patch clamp to the α1β2γ2L may affect the effect of anesthetics and induce different effects and α1β2γ2S GABAA receptors that are expressed on human from subject to subject. embryonic kidney (HEK) 293 cells with using midazolam and/ Midazolam is a popular sedative and sevoflurane is a most or sevoflurane. commonly used inhalational anesthetic. It is known that these drugs show their sedative, hypnotic or anesthetic actions Materials and Methods by binding to GABAA receptors. The GABAA receptor has the loci for binding to benzodiazepine, barbiturate and steroids. Transfection of the γ2 cDNA of the GABAA receptor to Midazolam activates the GABAA receptor by augmenting the HEK cells the binding of GABA to the receptor and midazolam directly activates the GABAA receptor at a high concentration. The γ2 HEK 293 cells are suitable for the transient expression of subunit is essential for binding benzodiazepine to the GABAA GABAA receptors and electrophysiological study [16]. Culturing 110 www.ekja.org Korean J Anesthesiol Eom, et al. of the HEK 293 cells and expressing GABAA cDNA in the HEK 1.5 mm, Harvard Apparatus Ltd., Edenbridge, Kent, UK). The 293 cells were performed as previously described [17]. resistance of the patch pipette was 4-6 M when it was filled The HEK 293 cells (CRL­1573; American Type Culture with intracellular solution (145 mM N­methyl­D­glucamine Collection) were cultured on glass cover slips in a solution HCl, 5 mM HEPES/KOH and 0.1 mM CaCl2, pH 7.2). containing minimum essential medium (MEM) supplemented The expression of GABA cDNA was confirmed by the develop­ with 10% heated fetal bovine serum, glutamine (4 mM), ment of intrinsic current, which was measured from the penicillin G (100 U/ml) and streptomycin sulfate (100 μg/ml). fluorescing HEK 293 cells, but not from the non­fluorescing Human γ2S cDNA was made from human γ2L cDNA (JC cells by the application of GABA (10 μM) and inhibition of the Biotech, Seoul, Korea) by deletion mutagenesis and its intrinsic current by the application of bicuculline (10 μM), sequence was confirmed by polymerase chain reaction. which is an antagonist of the GABAA receptor. The γ­aminobutyric acid type A receptor complementary In addition to the continuous bath perfusion with extracellular DNAs (the human γ2L or γ2S cDNA, the rat α1 cDNAs and the solution, solutions including GABA, sevoflurane or midazolam rat β2 cDNAs: the rat α1 and β2 cDNAs were generous gifts from were applied to the cells for 3 s at a rate of 10 ml/min using a Dr.
Load more

Recommended publications
  • Amnestic Concentrations of Sevoflurane Inhibit Synaptic
    Anesthesiology 2008; 108:447–56 Copyright © 2008, the American Society of Anesthesiologists, Inc. Lippincott Williams & Wilkins, Inc. Amnestic Concentrations of Sevoflurane Inhibit Synaptic Plasticity of Hippocampal CA1 Neurons through ␥-Aminobutyric Acid–mediated Mechanisms Junko Ishizeki, M.D.,* Koichi Nishikawa, M.D., Ph.D.,† Kazuhiro Kubo, M.D.,‡ Shigeru Saito, M.D., Ph.D.,§ Fumio Goto, M.D., Ph.D.࿣ Background: The cellular mechanisms of anesthetic-induced for surgical procedures do not have recollection of ac- amnesia are still poorly understood. The current study exam- tually being awake despite being awake and cooperative ined sevoflurane at various concentrations in the CA1 region of during the procedure.1 Galinkin et al.2 compared sub- rat hippocampal slices for effects on excitatory synaptic trans- Downloaded from http://pubs.asahq.org/anesthesiology/article-pdf/108/3/447/366512/0000542-200803000-00017.pdf by guest on 29 September 2021 mission and on long-term potentiation (LTP), as a possible jective, psychomotor, cognitive, and analgesic effects of mechanism contributing to anesthetic-induced loss of recall. sevoflurane (0.3% and 0.6%) with those of nitrous oxide Methods: Population spikes and field excitatory postsynaptic at equal minimum alveolar concentrations (MACs) in potentials were recorded using extracellular electrodes after healthy volunteers. They found that sevoflurane pro- electrical stimulation of Schaffer-collateral-commissural fiber inputs. Paired pulse facilitation was used as a measure of pre- duced a greater degree of amnesia and psychomotor synaptic effects of the anesthetic. LTP was induced using tetanic impairment than did an equal MAC of nitrous oxide but stimulation (100 Hz, 1 s). Sevoflurane at concentrations from had no analgesic actions.
    [Show full text]
  • Picrotoxin-Like Channel Blockers of GABAA Receptors
    COMMENTARY Picrotoxin-like channel blockers of GABAA receptors Richard W. Olsen* Department of Molecular and Medical Pharmacology, Geffen School of Medicine, University of California, Los Angeles, CA 90095-1735 icrotoxin (PTX) is the prototypic vous system. Instead of an acetylcholine antagonist of GABAA receptors (ACh) target, the cage convulsants are (GABARs), the primary media- noncompetitive GABAR antagonists act- tors of inhibitory neurotransmis- ing at the PTX site: they inhibit GABAR Psion (rapid and tonic) in the nervous currents and synapses in mammalian neu- system. Picrotoxinin (Fig. 1A), the active rons and inhibit [3H]dihydropicrotoxinin ingredient in this plant convulsant, struc- binding to GABAR sites in brain mem- turally does not resemble GABA, a sim- branes (7, 9). A potent example, t-butyl ple, small amino acid, but it is a polycylic bicyclophosphorothionate, is a major re- compound with no nitrogen atom. The search tool used to assay GABARs by compound somehow prevents ion flow radio-ligand binding (10). through the chloride channel activated by This drug target appears to be the site GABA in the GABAR, a member of the of action of the experimental convulsant cys-loop, ligand-gated ion channel super- pentylenetetrazol (1, 4) and numerous family. Unlike the competitive GABAR polychlorinated hydrocarbon insecticides, antagonist bicuculline, PTX is clearly a including dieldrin, lindane, and fipronil, noncompetitive antagonist (NCA), acting compounds that have been applied in not at the GABA recognition site but per- huge amounts to the environment with haps within the ion channel. Thus PTX major agricultural economic impact (2). ͞ appears to be an excellent example of al- Some of the other potent toxicants insec- losteric modulation, which is extremely ticides were also radiolabeled and used to important in protein function in general characterize receptor action, allowing and especially for GABAR (1).
    [Show full text]
  • GABA Receptors
    D Reviews • BIOTREND Reviews • BIOTREND Reviews • BIOTREND Reviews • BIOTREND Reviews Review No.7 / 1-2011 GABA receptors Wolfgang Froestl , CNS & Chemistry Expert, AC Immune SA, PSE Building B - EPFL, CH-1015 Lausanne, Phone: +41 21 693 91 43, FAX: +41 21 693 91 20, E-mail: [email protected] GABA Activation of the GABA A receptor leads to an influx of chloride GABA ( -aminobutyric acid; Figure 1) is the most important and ions and to a hyperpolarization of the membrane. 16 subunits with γ most abundant inhibitory neurotransmitter in the mammalian molecular weights between 50 and 65 kD have been identified brain 1,2 , where it was first discovered in 1950 3-5 . It is a small achiral so far, 6 subunits, 3 subunits, 3 subunits, and the , , α β γ δ ε θ molecule with molecular weight of 103 g/mol and high water solu - and subunits 8,9 . π bility. At 25°C one gram of water can dissolve 1.3 grams of GABA. 2 Such a hydrophilic molecule (log P = -2.13, PSA = 63.3 Å ) cannot In the meantime all GABA A receptor binding sites have been eluci - cross the blood brain barrier. It is produced in the brain by decarb- dated in great detail. The GABA site is located at the interface oxylation of L-glutamic acid by the enzyme glutamic acid decarb- between and subunits. Benzodiazepines interact with subunit α β oxylase (GAD, EC 4.1.1.15). It is a neutral amino acid with pK = combinations ( ) ( ) , which is the most abundant combi - 1 α1 2 β2 2 γ2 4.23 and pK = 10.43.
    [Show full text]
  • Exploring the Activity of an Inhibitory Neurosteroid at GABAA Receptors
    1 Exploring the activity of an inhibitory neurosteroid at GABAA receptors Sandra Seljeset A thesis submitted to University College London for the Degree of Doctor of Philosophy November 2016 Department of Neuroscience, Physiology and Pharmacology University College London Gower Street WC1E 6BT 2 Declaration I, Sandra Seljeset, confirm that the work presented in this thesis is my own. Where information has been derived from other sources, I can confirm that this has been indicated in the thesis. 3 Abstract The GABAA receptor is the main mediator of inhibitory neurotransmission in the central nervous system. Its activity is regulated by various endogenous molecules that act either by directly modulating the receptor or by affecting the presynaptic release of GABA. Neurosteroids are an important class of endogenous modulators, and can either potentiate or inhibit GABAA receptor function. Whereas the binding site and physiological roles of the potentiating neurosteroids are well characterised, less is known about the role of inhibitory neurosteroids in modulating GABAA receptors. Using hippocampal cultures and recombinant GABAA receptors expressed in HEK cells, the binding and functional profile of the inhibitory neurosteroid pregnenolone sulphate (PS) were studied using whole-cell patch-clamp recordings. In HEK cells, PS inhibited steady-state GABA currents more than peak currents. Receptor subtype selectivity was minimal, except that the ρ1 receptor was largely insensitive. PS showed state-dependence but little voltage-sensitivity and did not compete with the open-channel blocker picrotoxinin for binding, suggesting that the channel pore is an unlikely binding site. By using ρ1-α1/β2/γ2L receptor chimeras and point mutations, the binding site for PS was probed.
    [Show full text]
  • Toxicological and Pharmacological Profile of Amanita Muscaria (L.) Lam
    Pharmacia 67(4): 317–323 DOI 10.3897/pharmacia.67.e56112 Review Article Toxicological and pharmacological profile of Amanita muscaria (L.) Lam. – a new rising opportunity for biomedicine Maria Voynova1, Aleksandar Shkondrov2, Magdalena Kondeva-Burdina1, Ilina Krasteva2 1 Laboratory of Drug metabolism and drug toxicity, Department “Pharmacology, Pharmacotherapy and Toxicology”, Faculty of Pharmacy, Medical University of Sofia, Bulgaria 2 Department of Pharmacognosy, Faculty of Pharmacy, Medical University of Sofia, Bulgaria Corresponding author: Magdalena Kondeva-Burdina ([email protected]) Received 2 July 2020 ♦ Accepted 19 August 2020 ♦ Published 26 November 2020 Citation: Voynova M, Shkondrov A, Kondeva-Burdina M, Krasteva I (2020) Toxicological and pharmacological profile of Amanita muscaria (L.) Lam. – a new rising opportunity for biomedicine. Pharmacia 67(4): 317–323. https://doi.org/10.3897/pharmacia.67. e56112 Abstract Amanita muscaria, commonly known as fly agaric, is a basidiomycete. Its main psychoactive constituents are ibotenic acid and mus- cimol, both involved in ‘pantherina-muscaria’ poisoning syndrome. The rising pharmacological and toxicological interest based on lots of contradictive opinions concerning the use of Amanita muscaria extracts’ neuroprotective role against some neurodegenerative diseases such as Parkinson’s and Alzheimer’s, its potent role in the treatment of cerebral ischaemia and other socially significant health conditions gave the basis for this review. Facts about Amanita muscaria’s morphology, chemical content, toxicological and pharmacological characteristics and usage from ancient times to present-day’s opportunities in modern medicine are presented. Keywords Amanita muscaria, muscimol, ibotenic acid Introduction rica, the genus had an ancestral origin in the Siberian-Be- ringian region in the Tertiary period (Geml et al.
    [Show full text]
  • Bicuculline and Gabazine Are Allosteric Inhibitors of Channel Opening of the GABAA Receptor
    The Journal of Neuroscience, January 15, 1997, 17(2):625–634 Bicuculline and Gabazine Are Allosteric Inhibitors of Channel Opening of the GABAA Receptor Shinya Ueno,1 John Bracamontes,1 Chuck Zorumski,2 David S. Weiss,3 and Joe Henry Steinbach1 Departments of 1Anesthesiology and 2Psychiatry, Washington University School of Medicine, St. Louis, Missouri 63110, and 3University of Alabama at Birmingham, Neurobiology Research Center and Department of Physiology and Biophysics, Birmingham, Alabama 35294-0021 Anesthetic drugs are known to interact with GABAA receptors, bicuculline only partially blocked responses to pentobarbital. both to potentiate the effects of low concentrations of GABA and These observations indicate that the blockers do not compete to directly gate open the ion channel in the absence of GABA; with alphaxalone or pentobarbital for a single class of sites on the however, the site(s) involved in direct gating by these drugs is not GABAA receptor. Finally, at receptors containing a1b2(Y157S)g2L known. We have studied the ability of alphaxalone (an anesthetic subunits, both bicuculline and gabazine showed weak agonist steroid) and pentobarbital (an anesthetic barbiturate) to directly activity and actually potentiated responses to alphaxalone. These activate recombinant GABAA receptors containing the a1, b2, and observations indicate that the blocking drugs can produce allo- g2L subunits. Steroid gating was not affected when either of two steric changes in GABAA receptors, at least those containing this mutated b2 subunits [b2(Y157S) and b2(Y205S)] are incorporated mutated b2 subunit. We conclude that the sites for binding ste- into the receptors, although these subunits greatly reduce the roids and barbiturates do not overlap with the GABA-binding site.
    [Show full text]
  • The Protective Effect of Nicardipine on Iron-Induced Purkinje
    Fırat Tıp Dergisi 2008;13(3): 167-170 Experimental Research www.firattipdergisi.com The Protective Effect of Nicardipine on Iron-Induced Purkinje Cell Loss in Rat Cerebellum: A Stereological Study Ramazan KOZAN a1 , M. Ömer BOSTANCI 2, Fatih SEFĐL 2, Faruk BAĞIRICI 2 1 Mustafa Kemal University, Faculty of Medicine, Department of Physiology, HATAY 2 Ondokuz Mayis University, Faculty of Medicine, Department of Physiology, SAMSUN ABSTRACT Objective: The aim of the present study is to investigate the effect of nicardipine, a calcium channel blocker, on the neurotoxicity induced by intracerebroventricular (i.c.v.) iron injection in rats. Materials and Methods: Animals were divided into three groups; control, iron and iron+nicardipine groups. Rats in iron and iron+nicardipine groups received i.c.v. FeCl 3, while rats in control group received the same volume of saline. All animals were kept alive for ten days following the operation and animals in iron+nicardipine group were injected intraperitoneally nicardipine (10 mg/kg/day) once a day during this period. After ten days, all rats were perfused intracardially and cerebellar tissues were stained with Cresyl violet. Means of total Purkinje cells numbers in the cerebellum were estimated using the optical fractionator counting method. Results: Means of total Purkinje cells numbers in the cerebellum as follows: 317182±9667, 209002±7836 and 265659±8291 in the control, iron and iron+nicardipine groups, respectively. Total number of Purkinje cells in iron and iron+nicardipine groups were significantly lower than control animals (p< 0.05). However, comparison between iron and iron+nicardipine groups revealed that nicardipine significantly attenuates the iron-induced Purkinje cell loss (p<0.05).
    [Show full text]
  • Molecular Changes in Opioid Addiction: the Role of Adenylyl Cyclase and Camp/PKA System 205
    CHAPTER SEVEN Molecular Changes in Opioid Addiction: The Role of Adenylyl Cyclase and cAMP/PKA System ,1 † Patrick Chan* , Kabirullah Lutfy * Department of Pharmacy and Pharmacy Administration, Western University of Health Sciences, College of Pharmacy, Pomona, California, USA † Department of Pharmaceutical Sciences, College of Pharmacy, Western University of Health Sciences, Pomona, California, USA 1 Corresponding author: e-mail address: [email protected]. Contents 1. Introduction 204 2. The Adenylyl Cyclase Pathway 205 2.1 Adenylyl Cyclase 205 2.2 Protein Kinase A 207 3. Opioid Effect on cAMP-Responsive Element-Binding Protein 209 4. Molecular Changes in Brain Regions That May Underlie Opiate Dependence 211 4.1 Molecular Changes in the Locus Coeruleus 211 4.2 Molecular Changes in the Amygdala 214 4.3 Molecular Changes in the Periaqueductal Gray 216 5. Molecular Changes in the Ventral Tegmental Area 217 6. Molecular Changes in Other CNS Regions 218 7. Conclusions 219 References 219 Abstract For centuries, opiate analgesics have had a considerable presence in the treatment of moderate to severe pain. While effective in providing analgesia, opiates are notorious in exerting many undesirable adverse reactions. The receptor targets and the intra- cellular effectors of opioids have largely been identified. Furthermore, much of the mechanisms underlying the development of tolerance, dependence, and withdrawal have been delineated. Thus, there is a focus on developing novel compounds or strategies in mitigating or avoiding the development of tolerance, dependence, and withdrawal. This review focuses on the adenylyl cyclase and cyclic adenosine 3,5-monophosphate (cAMP)/protein kinase A (AC/cAMP/PKA) system as the central player in mediating the acute and chronic effects of opioids.
    [Show full text]
  • Effects of Gabaa-Active Agents on Thermoregulataion in Rats
    Trakia Journal of Sciences, Vol. 8, Suppl. 2, pp 102-106, 2010 Copyright © 2009 Trakia University Available online at: http://www.uni-sz.bg ISSN 1313-7050 (print) ISSN 1313-3551 (online) EFFECTS OF GABAA-ACTIVE AGENTS ON THERMOREGULATAION IN RATS R. Nikolov*, K. Yakimova Department of Pharmacology and Toxicology, Medical Faculty, Medical University, Sofia ABSTRACT PURPOSE: Gamma-aminobutyric acid (GABA) is the principal inhibitory neurotransmitter, which is widely distributed throughout the mammalian brain including hypothalamus. Immunohistochemical research have reported GABA-ergic neurons and GABAA-receptors on the neurons of the preoptic area of anterior hypothalamus (PO/AH). The aim of this study was to investigate the influence of GABAA- ergic substances on thermoregulation in rats. METHODS: We have studied the effects of GABAA- active agents, muscimol and diazepam on core body temperature in rats after systemic administration (intraperitoneally, i.p.). Body temperature was measured with thermistor probes (TX8) and monitored on multichannel recorder THERMEX 16. RESULTS: Intraperitoneal injection of muscimol or diazepam has produced dose-dependent hypothermia. Hypothermic effect of muscimol was inhibited by pretreatment of bicuculline, a competitive antagonist of GABAA-receptors. Diazepam induced hypothermia was antagonized by pretreatment of animals with flumazenil, a competitive antagonist of benzodiazepine receptors. CONCLUSION: Hypothermia induced by muscimol or diazepam suggest involvement of GABAA receptors in the processes
    [Show full text]
  • Opposing Actions of Sevoflurane on Gabaergic and Glycinergic Synaptic Inhibition in the Spinal Ventral Horn
    Opposing Actions of Sevoflurane on GABAergic and Glycinergic Synaptic Inhibition in the Spinal Ventral Horn Veit-Simon Eckle, Sabrina Hauser, Berthold Drexler*, Bernd Antkowiak, Christian Grasshoff Experimental Anesthesiology Section, Department of Anesthesiology & Intensive Care, Tu¨bingen University Hospital, Eberhard-Karls-University, Tu¨bingen, Germany Abstract Background: The ventral horn is a major substrate in mediating the immobilizing properties of the volatile anesthetic sevoflurane in the spinal cord. In this neuronal network, action potential firing is controlled by GABAA and glycine receptors. Both types of ion channels are sensitive to volatile anesthetics, but their role in mediating anesthetic-induced inhibition of spinal locomotor networks is not fully understood. Methodology/Principal Findings: To compare the effects of sevoflurane on GABAergic and glycinergic inhibitory postsynaptic currents (IPSCs) whole-cell voltage-clamp recordings from ventral horn interneurons were carried out in organotypic spinal cultures. At concentrations close to MAC (minimum alveolar concentration), decay times of both types of IPSCs were significantly prolonged. However, at 1.5 MAC equivalents, GABAergic IPSCs were decreased in amplitude and reduced in frequency. These effects counteracted the prolongation of the decay time, thereby decreasing the time- averaged GABAergic inhibition. In contrast, amplitudes and frequency of glycinergic IPSCs were not significantly altered by sevoflurane. Furthermore, selective GABAA and glycine receptor antagonists were tested for their potency to reverse sevoflurane-induced inhibition of spontaneous action potential firing in the ventral horn. These experiments confirmed a weak impact of GABAA receptors and a prominent role of glycine receptors at a high sevoflurane concentration. Conclusions: At high concentrations, sevoflurane mediates neuronal inhibition in the spinal ventral horn primarily via glycine receptors, and less via GABAA receptors.
    [Show full text]
  • Benzamil Inhibits Neuronal and Heterologously Expressed Small
    *Manuscript Click here to view linked References Benzamil inhibits neuronal and heterologously expressed small conductance Ca2+-activated K+ channels Marisol Sampedro Castañeda1,3, Raffaella Tonini1,2 , Christopher D. Richards1, Martin Stocker1* and Paola Pedarzani1* 1Research Department of Neuroscience, Physiology and Pharmacology, University College London, London, UK. 2Neuromodulation of Cortical and Subcortical Circuits Laboratory, Fondazione Istituto Italiano di Tecnologia, Genova, Italy. 3Kinases and Brain Development Laboratory, The Francis Crick Institute, London, UK ORCID: Paola Pedarzani - https://orcid.org/0000-0002-2665-7911 Martin Stocker - https://orcid.org/0000-0003-3993-5666 *To whom correspondence should be addressed at: Research Department of Neuroscience, Physiology and Pharmacology University College London, Gower Street, London WC1E 6BT, United Kingdom Tel.: +44-(0)20-7679-7744 (PP) or +44-(0)20-3549-5671(MS) E-mail: [email protected] or [email protected] 1 Abstract Small conductance Ca2+-activated K+ (SK) channels are expressed throughout the soma and dendrites of pyramidal neurons in the neocortex and hippocampal formation, where they participate in the local regulation of membrane excitability and synaptic signals. Through their inter-play with Ca2+ channels, SK channels regulate Ca2+ influx triggered by back-propagating action potentials in dendrites. Inhibition of SK channels affects both the amplitude and duration of Ca2+ transients, but the role of Ca2+ clearance mechanisms and their link to SK channel activity has not been established. Here we report the effect of the Na+/Ca2+ exchanger (NCX) inhibitor benzamil on Ca2+ extrusion and SK channels in the regulation of dendritic Ca2+ signals. Benzamil increased the duration and amplitude of dendritic Ca2+ transients elicited by back- propagating action potentials in hippocampal pyramidal neurons.
    [Show full text]
  • A Molecular Mechanism for Choosing Alcohol Over an Alternative Reward Eric Augier, Estelle Barbier, Russell S
    Corrected 9 July 2018. See full text. RESEARCH ALCOHOL DEPENDENCY ward 26.2% of the time (Fig. 1C). However, al- though the vast majority of rats strongly preferred saccharin,aminority,4ratsoutof32inthefirst A molecular mechanism for choosing experiment (12.5% of the population, Fig. 1D, left) continued to choose alcohol despite having access to a high-value alternative (alcohol-preferring, alcohol over an alternative reward AP). Although this was a small number, it aligns well with human addiction rates (7, 8) and promp- Eric Augier1*, Estelle Barbier1, Russell S. Dulman2, Valentina Licheri3, Gaëlle Augier1, ted us to expand the study of individual differences Esi Domi1, Riccardo Barchiesi1, Sean Farris4, Daniel Nätt1, R. Dayne Mayfield4, in choice behavior. Ultimately, this percentage Louise Adermark3, Markus Heilig1 was stable across a large number of rats from successive batches (see below; Fig. 1D, right). Alcohol addiction leads to increased choice of alcohol over healthy rewards. We established Alcohol preference was not influenced by hold- an exclusive choice procedure in which ~15% of outbred rats chose alcohol over a ing prior history of alcohol and saccharin self- high-value reward. These animals displayed addiction-like traits, including high motivation administration identical (fig. S3). to obtain alcohol and pursuit of this drug despite adverse consequences. Expression of the g-aminobutyric acid (GABA) transporter GAT-3 was selectively decreased within the Extensive pre-exposure to saccharin amygdala of alcohol-choosing rats, whereas a knockdown of this transcript reversed choice does not affect subsequent preference of rats that originally chose a sweet solution over alcohol. GAT-3 expression alcohol choice was selectively decreased in the central amygdala of alcohol-dependent people compared People who go on to develop addictive disorders to those who died of unrelated causes.
    [Show full text]