Comparison of GABA analogues at the crayfish stretch receptor neurone (original) (raw)
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Neuroscience Letters, 1984
The conductance increase induced by bath application of GABA has been measured in voltageclamped stretch-receptor neurones of crayfish. A rapid conductance increase was obtained only at GABA concentrations above 3.3 x 10-4 M. The response to lower GABA concentrations (between 10-4 and 10-6 M) developed slowly over 30-60 min. Repetitive application of intermediate GABA concentrations induced postsynaptic conductance changes which were progressively enhanced in their onset and magnitude. In the presence of nipecotic acid or in Na +-free Ringer solutions, the response to all GABA concentrations was rapid and constant for each concentration. The time course of inhibitory postsynaptic currents was unaffected by nipecotic acid. These results suggest the presence of a saturable GABA uptake system which limits the access of bath-applied GABA to postsynaptic receptors. This system has little if any effect on the termination of response to synaptically released GABA.
Inward current caused by sodium-dependent uptake of GABA in the crayfish stretch receptor neurone
The Journal of Physiology, 1992
1. A two-microelectrode current-voltage clamp and Cl-selective microelectrodes were used to examine the effects of y-aminobutyric acid (GABA) on membrane potential, current and intracellular Clactivity (a 1) in the crayfish stretch receptor neurone. All experimental solutions were CO2-HC03free. 2. GABA (500 ftM) produced a monoor biphasic depolarization (amplitude < 10 mV), often with a prominent initial depolarizing component followed by a transient shift to a more negative level. In some neurones, an additional depolarizing phase was seen upon washout of GABA. Receptor desensitization, being absent, played no role in the multiphasic actions of GABA. 3. The pronounced increase in membrane conductance evoked by GABA (500 /tM) was associated with an increase in a'1 which indicates that the depolarizing action was not due to a current carried by Clions. 4. The currents activated by GABA under voltage clamp conditions were inwardly directed when recorded at the level of the resting membrane potential, and they often revealed a biphasic character. The reversal potential of peak currents activated by pulses of 500 jtM-GABA (EGABA) was 9-12 mV more positive than the reversal potential of the simultaneously measured net Clflux (Ec1). Ec1 was 2-7 mV more negative than the resting membrane potential. 5. EGABA (measured using pulses of 500 4M-GABA) was about 10 mV more positive than the reversal potential of the current activated by 500 /tM-muscimol, a GABA agonist that is a poor substrate of the Na+-dependent GABA uptake system. 6. In the absence of Na', the depolarization and inward current caused by 500 pM-GABA were converted to a hyperpolarization and to an outward current. Muscimol produced an immediate outward current both in the presence and absence of Na'. 7. Following block of the inhibitory channels by picrotoxin (100-200 ftM), the depolarizing effect of 500 ,tM-GABA was enhanced and the transient hyperpolarizing shifts were abolished. 8. In the presence of picrotoxin, GABA (>2 aM) produced a concentrationdependent monophasic inward current which had a reversal potential of +30 to + 60 mV. This current was inhibited in the absence of Na' and by the GABA uptake blocker, nipecotic acid. Unlike the channel-mediated current, the picrotoxin-MS 9386 628 K. KAILA, B. RYDQVIST, M. PASTERNACK AND J. VOIPJO insensitive current was activated without delay also at low (2-10 Am) concentrations of GABA. 9. Brief (< 10 s) pulses of GABA at a low concentration (< 100 sam) produced only a small increase in conductance, and the reversal potential of the GABA-activated current obtained in this manner was close to that seen in the presence of picrotoxin. In contrast to this, the reversal potential of the current activated by pulses of 50 pMmuscimol was identical to that observed at 500 /m. 10. The present results indicate that a sodium-dependent electrogenic GABA uptake mechanism has a direct influence on the current and voltage responses evoked by GABA in the crayfish stretch receptor neurone. The current component attributable to uptake makes EGABA significantly more positive than E(> and explains the mono-and biphasic depolarizing actions of GABA. When measured using low concentrations and short pulses of GABA, the preferential activation of the uptake mechanism leads to an estimate of EG ABA which is close to the reversal potential of the uptake current.
The Journal of Physiology, 1981
1. The effect of pentobarbitone (PB) on GABA-ergic inhibition was investigated in the isolated crayfish stretch receptor. The soma of the slowly adapting neurone was impaled with two micro-electrodes to give an accurate determination of membrane conductances. 2. Application of PB in concentrations from 106 to 10-M increased the rise time constant of the inhibitory pQst-synaptic potential (i.p.s.p.). The i.p.s.p. percentage amplitude and decay time constant were also increased in eight out of twelve neurones. On prolonged exposure, the percentage amplitude declined at a rate dependent upon the dose and the frequency of stimulation until the i.p.s.p. became undetectable. 3. The response to ionophoretically applied GABA remained essentially unaltered in the presence of PB, but the falling phase was prolonged by up to 8 % in four of the ten neurones tested. Resting membrane conductance, i.p.s.p. driving force (i.p.s.p. reversal potential minus resting membrane potential), and parameters of the antiand orthodromic action potential were not significantly affected. 4. Removal of PB after prolonged exposure usually caused an immediate increase in i.p.s.p. percentage amplitude but the i.p.s.p. rising phase remained slowed. 5. Application of excess extracellular GABA only affected the i.p.s.p. percentage amplitude after it had been reduced by PB. It transiently increased the attenuated i.p.s.p. percentage amplitude in the presence of PB, and after the removal of PB permanently increased the amplitude to its original value. 6. Nipecotic acid and cis-1,3-aminocyclohexane carboxylic acid, inhibitors of GABA re-uptake, slightly increased the i.p.s.p. percentage amplitude, and prolonged the falling phase but did not affect the rising phase. The percentage amplitude declined on prolonged exposure. 7. We conclude that PB has no electrophysiologically demonstrable post-synaptic action in the crayfish stretch receptor neurone, but it inhibits the presynaptic release and re-uptake of GABA.
Crayfish sensory terminals and motor neurones exhibit two distinct types of GABA receptors
Journal of Comparative Physiology A: Sensory, Neural, and Behavioral Physiology, 1996
Motor neurones of the crayfish walking system display inhibitory responses evoked either byamino butyric acid (GABA) or glutamate, possibly involving the same receptor (Pearlstein et al. 1994). In order to test if this sensibility to both GABA and glutamate was a specific property of crayfish GABA receptors, pharmacological characteristics of GABAevoked responses in both sensory terminals from CB chordotonal organ and motor neurones of the walking system have been compared. Both receptors are GABAgated Cl) channels activated by specific GABA A (muscimol, isoguvacine), GABA B (3-aminopropyl phosphinic acid), and GABA C (cis-4-amino crotonic acid) agonists, and blocked by competitive (-guanidino propionic acid) and non-competitive (picrotoxin) antagonists. They were insensitive to specific GABA A (bicuculline, SR-95531) and GABA B (phaclofen) antagonists. Furthermore, in both cases, nipecotic acid and the modulatory drug diazepam had no effect. However, our results demonstrate that GABA receptors of sensory terminals are different from those of motor neurones. GABA-induced desensitisation only occurred in sensory terminals. Moreover, glutamate was shown to activate GABA-gated Cl) channels in motor neurones, but not in sensory terminals. Therefore, GABA is likely to be the endogenous neurotransmitter of presynaptic inhibition in sensory terminals, whereas inhibition between antagonistic motor neurones would be achieved by glutamate. Key words GABA receptors • Glutamate • Sensory terminals • Motor neurones • Invertebrate Abbreviations 3-APPA 3-aminopropyl phosphinic acid • 3-APA 3-aminopropyl phosphonous acid • CACA cis-4-amino crotonic acid • CBCO coxo-basipodite chordotonal organ • GABA-amino butyric acid •-GP-guanidinopropionic acid • PTX picrotoxin • R i input resistance • T e ejection time • V m membrane potential • V r resting potential
An intracellular study of the effects of GABA on frog tectal neurones in vitro
Neuroscience Letters, 1992
The effects of y-aminobutyric acid (GABA) on neurones of the amphibian optic tectum were studied with current-and voltage-clamp recording from an isolated preparation of the midbrain of the frog Rana temporaria. Bath-applied GABA (1 mM) enhanced depolarizing synaptic potentials evoked in layer 6 tectal neurones by orthodromic stimulation of the optic tract. GABA also facilitated Na*-and Ca2+-dependent action potentials elicited by intracellular injection of depolarizing current. These actions of GABA were associated with comparatively small changes in membrane potential and their reversal potential was dependent on the CI equilibrium potential. Changes in input resistance observed during application of GABA were small and in part accountable for by the rectifying properties of the cell membrane. Tetrodotoxin (TTX; 1/.tM) did not block the action of GABA on these neurones. These results show that externally applied GABA was able to raise directly the intrinsic excitability of frog tectal neurones and to enhance excitatory synaptic transmission elicited by stimulation of optic nerve fibres.
Effect of gamma-aminobutyric acid on intracellular pH in the crayfish stretch-receptor neurone
Journal of Experimental Biology, 1991
The effect of gamma-aminobutyric acid (GABA) on intracellular pH (pHi) was examined in the crayfish stretch-receptor neurone using H(+)-selective microelectrodes and a two-microelectrode voltage clamp. In the presence of 30 mmol l-1 HCO3- (pH 7.4), application of GABA (0.5 mmol l-1) produced a mean fall in pHi of 0.26 units. The initial rate of fall of pHi was attributable to a net influx of acid equivalents of 6.3 mmol l-1 min-1. In the nominal absence of HCO3-, GABA had little effect on pHi. The HCO3(−)-dependent acidosis caused by GABA was inhibited by picrotoxin (0.1 mmol l-1) but not by depletion of extracellular and intracellular Cl-. Acetazolamide (0.1 mmol l-1) decreased the rate of fall of pHi caused by a step increase in CO2 partial pressure as well as by GABA, which indicates that the neurone contains carbonic anhydrase. In the presence of both Cl- and HCO3-, the reversal potential of the GABA-activated current was more positive than under nominally HCO3(−)-free condition...
The Journal of Neuroscience, 1981
The structure of the GABA receptor was investigated by determining the relative effects of two stereoisomers of the GABA agonist, y-amino-/&hydroxybutyric acid (GABOB), in several quantitative receptor-related assay systems. (3S)-(+)-4-Amino-3-hydroxybutanoic acid (d-GABOB) was found to be about twice as potent as (3R)-(-)-4-amino-3-hydroxybutanoic acid (I-GABOB) in displacing ["Hlmuscimol from specific binding sites in mouse brain membrane fractions. ["H]Muscimol is thought to bind to the GABA recognition site of postsynaptic GABA receptor.anionophore complexes. A similar order of potency for the GABOB enantiomers was observed for the cerebrovascular GABA receptor in ["Hlmuscimol binding assays using bovine cerebral blood vessels. In contrast to the binding results, I-GABOB was significantly more potent than d-GABOB in mimicking the postsynaptic action of GABA, which was measured as increases in membrane input conductance in the isolated crayfish stretch receptor neuron. Both GABOB enantiomers have some affinity for GABA transport processes, and d-GABOB was found to be more potent than I-GABOB in inhibiting GABA uptake into rat brain synaptosomes and Na'-dependent GABA binding to mouse brain membranes. I-GABOB was more potent than d-GABOB when conductance measurements were made in the presence of lo-" M nipecotic acid or L-a&diaminopropionic acid, two specific GABA transport blockers. The greater effectiveness ' This work was supported by United States Public Health Service Grants NS 12116 and 01615. We wish to thank Dr. K. Ikeda for instruction and guidance with the crayfiih stretch receptor assay, Dr. Y.-J. Wang for furnishing data dealing with membrane detergent treatments, and Miss Phyllis Degener for invaluable technical assistance. procedures to measure GABA binding in brain membrane preparations to presumed GABA recognition sites of receptor. anionophore complexes (Enna and Snyder, 1975) and of the GABA transport system (Roberts et al., 197813) and to GABA receptor sites in cerebral blood vessels (Krause et al., 1980). In general, we found a good quantitative correspondence between the physiological and biochemical measurements (Roberts et al., 1978a; Krause et al., 1980; D. N. Krause, K. Ikeda, and E. Roberts, manuscript in preparation). In the course of the above studies, we investigated the relative effects of two stereoisomers of the GABA agonist y-amino-fi-hydroxybutyric acid (GABOB), (3S)-(+)-4amino-3-hydroxybutanoic acid (d-GABOB) and (3R)-(-)-4-amino-3-hydroxybutanoic acid (I-GABOB). Much to our surprise, the results of the present investigation showed that d-GABOB was more potent than I-GABOB in in vitro binding and uptake systems, while the reverse was found to be true in our conductance studies with the crayfish stretch receptor neuron and in the physiological
Effect of γ-Aminobutyric Acid on Intracellular pH in the Crayfish Stretch-Receptor Neurone
Journal of Experimental Biology, 1991
The effect of γ-aminobutyric acid (GABA) on intracellular pH (pHi) was examined in the crayfish stretch-receptor neurone using H+-selective microelectrodes and a two-microelectrode voltage clamp. In the presence of 30 mmol I−1 HCO3− (pH 7.4), application of GABA (0.5 mmol I−1) produced a mean fall in pHi of 0.26 units. The initial rate of fall of pHi was attributable to a net influx of acid equivalents of 6.3 mmol I−1 min−1. In the nominal absence of HCO3−, GABA had little effect on pHi. The HCO3−-dependent acidosis caused by GABA was inhibited by picrotoxin (0.1 mmol I−1) but not by depletion of extracellular and intracellular Cl−. Acetazolamide (0.1 mmol I−1) decreased the rate of fall of pHi caused by a step increase in CO2 partial pressure as well as by GABA, which indicates that the neurone contains carbonic anhydrase. In the presence of both Cl− and HCO3−, the reversal potential of the GABA-activated current was more positive than under nominally HCO3−-free conditions. In line...
EFFECT OF y-AMINOBUTYRIC ACID ON EXTRACELLULAR pH IN THE CRAYFISH STRETCH-RECEPTOR NEURONE
1991
The effect of y-aminobutyric acid (GABA) on intracellular pH (pHi) was examined in the crayfish stretch-receptor neurone using H +-selective microelectrodes and a two-microelectrode voltage clamp. In the presence of SOmmoll" 1 HCO 3~ (pH7.4), application of GABA (O.Smmoll" 1) produced a mean fall in pHi of 0.26 units. The initial rate of fall of pHi was attributable to a net influx of acid equivalents of 6.3mmol I" 1 min" 1. In the nominal absence of HCO 3~, GABA had little effect on pHi. The HCO 3~-dependent acidosis caused by GABA was inhibited by picrotoxin (O.lmmoir 1) but not by depletion of extracellular and intracellular C\'. Acetazolamide (0.1 mmol I" 1) decreased the rate of fall of pHi caused by a step increase in CO 2 partial pressure as well as by GABA, which indicates that the neurone contains carbonic anhydrase. In the presence of both Cl~ and HCO 3 ", the reversal potential of the GABA-activated current was more positive than under nominally HCO 3~-free conditions. In line with this, GABA induced a marked HCO 3~-dependent depolarization, and this depolarizing action was enhanced in the absence of Cl~ so as to lead to triggering of action potentials. All these observations support the conclusion that the GABA-induced fall in pHi is due to a net efflux of HCO 3~ through the inhibitory anion channels.