Jack Benson - Academia.edu (original) (raw)
Papers by Jack Benson
Nature, 2000
The quality of Fig. 2 was unsatisfactory as published. The ®gure is reproduced again here. M α1 α... more The quality of Fig. 2 was unsatisfactory as published. The ®gure is reproduced again here. M α1 α2 α3 Wild-type α1(H101R) γ2 β2/3 α5 Wild-type α1(H101R) a Wild-type α1(H101R) 28 ± 15 Diazepam > 10,000 Clonazepam 3 ± 2 > 10,000 Zolpidem 13 ± 5 > 10,000 Displacement of [ 3 H]Ro15-4513, K i (nM) c Wild-type α1(H101R) α1 b α1(H101R) Wild-type d e Wild-type 3 µM GABA+Dz +Ro15 3 µM GABA α1(H101R) 3 µM GABA+Dz +Ro15 3 µM GABA 500 pA 2 s [ 3 H]Ro15-4513 [ 3 H]Ro15-4513, K i (nM) + diazepam
Journal of Experimental Biology, 1984
Dopamine, a cardioexcitor in decapod crustaceans, increased the frequency and/or duration of burs... more Dopamine, a cardioexcitor in decapod crustaceans, increased the frequency and/or duration of bursts of action potentials in the semi-isolated cardiac ganglia of two species of crabs. The number of motoneurone action potentials in each burst was increased, which in the intact heart would increase the force and amplitude of heart contraction. The effects were concentration-dependent, with a threshold concentration of 10−8M or lower when dopamine was applied by continuous perfusion. At 5 × 10−6M, dopamine increased burst frequency by 200%. The main site of dopamine action was the group of four posterior small interneurones which normally function as the pacemaker for the cardiac ganglion system. Effects on the five large motoneurones occurred at higher concentrations. This regional difference in sensitivity was demonstrated by selective applications of dopamine to different parts of the cardiac ganglion and by the use of preparations in which the two ends of the ganglion had been funct...
Journal of Experimental Biology, 1989
The isolated, intact heart of the marine arachnid Limulus polyphemus continues to beat in vitro f... more The isolated, intact heart of the marine arachnid Limulus polyphemus continues to beat in vitro for many hours. Application of γ-aminobutyric acid (GABA) decreased the heart beat frequency with a threshold of 3×10−7moll−1 and an EC50 of 2·0±0·6×10−5moll−1 (mean±S.D., N = 8). At 10−4moll−1 and above the heart beat was completely and reversibly inhibited. The agonist potency profile of the Limulus heart chronotropic GABA receptor was very similar to that of the vertebrate GABAA receptor: muscimol > ZAPA>GABA≈TACA>isoguvacine>THIP>3-aminopropane sulphonic acid> imidazole-4-acetic acid ≈β-guanidino proprionic acid ≈5-aminovalerate. In contrast, the antagonist profile differed dramatically: bicuculline, pitrazepin and SR 95103, as well as the channel blocker picrotoxin, were without effect at concentrations up to 10−4moll−1. The benzodiazepines clorazepate, flunitrazepam, flurazepam and diazepam, as well as the barbiturate sodium pentobarbital, were without effect on th...
Journal of Experimental Biology, 1980
The five large and four small neurones in the cardiac ganglion of the crab, Portunus, are electro... more The five large and four small neurones in the cardiac ganglion of the crab, Portunus, are electrotonically coupled and behave as a single relaxation oscillator, exhibiting periodic bursting activity in vitro. Recorded from the large neurone somata, this activity consists of 200–400 ms slow depolarizations called ‘driver potentials’ (Tazaki & Cooke, 1979a), accompanied by attenuated action potentials and EPSP’s from small neurone input. There is a strong positive correlation between the duration of the driver potential and the duration of the following interburst interval in the spontaneously active ganglion. This correlation is preserved during prolonged depolarization and hyperpolarization. When a driver potential is prematurely terminated by an injected current pulse, the following interburst interval is shortened in direct proportion to the decrease in driver potential duration. When a driver potential or a burst of high-frequency action potential activity is evoked by a depolari...
Journal of Experimental Biology, 1992
1. Three different responses were evoked by pressure micro-application of serotonin onto freshly ... more 1. Three different responses were evoked by pressure micro-application of serotonin onto freshly dissociated, current- and voltage-clamped neuronal somata from the thoracic ganglia of the locust Locusta migratoria. 2. In some neurones, an inward current, I(5HT)K, resulting from a decrease in potassium conductance, with slow kinetics and maximum activation at membrane potentials of −60 to - 70 mV, was evoked by serotonin and by the 5-HT3 agonist 2-methyl serotonin. This current was completely abolished by either 10 mmoll−1 caesium or 5 mmoll−1 rubidium and partially blocked by 50 mmoll−1 tetraethylammonium or 5 mmoll−1 4-aminopyridine. The response was antagonised by the 5-HT2-specific compounds, ketanserin and ritanserin. 3. In other somata, serotonin, 2-methyl serotonin and the 5-HT3 antagonist ICS205 930 evoked a second current, I(5HT)Na, which was due to an increase in sodium permeability and had slow kinetics similar to that of I(5HT)K. This current was inward over the membrane ...
Trends in Neurosciences, 1984
The organization into bursts of action potential firing is a characteristic of the neurons of man... more The organization into bursts of action potential firing is a characteristic of the neurons of many groups of animals. In the crustacean cardiac and stomatogastric ganglia, burst generation and perhaps also endogenous rhythmiciry result from the production of slow, &+-dependent, non-propagated depolarizations known as driver or plateau potentials. At least some neuromodulatory inputs to these ganglia have their effects by acting on these potentials, and driver potential formation appears to be an essential aspect of non-impulse-mediated synaptic coordination in these ganglia.
Proceedings of the National Academy of Sciences, 1983
Previous work has shown that serotonin causes an increase in K+ conductance in the identified Apl... more Previous work has shown that serotonin causes an increase in K+ conductance in the identified Aplysia neuron R15. This response is mediated by cAMP-dependent protein phosphorylation. The results presented here show that the K+ channel modulated by serotonin is an anomalous or inward rectifier (designated IR) that is present in R15 together with the three other distinct K+ channels previously described for this cell. Several lines of evidence indicate that this inward rectifier is partially activated in the resting cell and is further activated by serotonin. Voltage clamp analysis of resting and serotonin-evoked membrane currents at various external K+ concentrations shows that both currents have reversal potentials close to the potassium equilibrium potential, exhibit similar dependences in magnitude on external K+ concentration, and display marked anomalous rectification. The effects of particular monovalent and divalent cations are also similar on the resting and serotonin-evoked ...
European Journal of Pharmacology, 2001
The functional pharmacology of receptors composed of the chicken brain GABA receptor g4 subunit a... more The functional pharmacology of receptors composed of the chicken brain GABA receptor g4 subunit and the mammalian GABA A A receptor a 3 and b2 subunits was studied by heterologous expression in Xenopus laeÕis oocytes using the two electrode voltage-clamp technique. GABA-evoked currents had an EC of 180 " 30 mM. Responses were blocked by the competitive and non-competitive 50 GABA receptor antagonists, bicuculline methochloride and picrotoxin. Sodium pentobarbital reversibly potentiated the current A 2q Ž. several-fold, and Zn ions blocked the current with high potency IC s 20 mM. GABA-evoked currents were potentiated by the 50 benzodiazepine site full agonists flunitrazepam and triazolam and less by the partial agonists abecarnil and bretazenil. The inverse Ž. Ž. agonists methyl-b-carboline-3-carboxylate b-CCM and methyl 6,7-dimethoxy-4-ethyl-b-carboline-3-carboxylate DMCM reduced the current. However, the imidazobenzodiazepine Ro 15-4513, which acts as an inverse agonist at mammalian a xb y g2 GABA receptors A
Brain Research, 1988
The isolated somata of neurons from the thoracic ganglia of the locust, Locusta migratoria, respo... more The isolated somata of neurons from the thoracic ganglia of the locust, Locusta migratoria, respond to pressure microapplication of y-aminobutyric acid (GABA) and acetylcholine. The acetylcholine receptors fall into two groups, ACh t (activated by nicotine) and ACh 2 (activated by muscarine). The GABA receptor and the ACh I receptor differ in pharmacology from the known vertebrate receptors. The GABA receptor is insensitive to bicuculline and its salts up to a concentration of 10-4 M. In contrast, bicuculline is a moderately potent, at least partially competitive antagonist of the ACh~ receptor-mediated response in the thoracic neuronal somata. These observations suggest that classical diagnostic compounds such as bicuculline may show greater cross-reactivity than hitherto suspected among the members of the superfamily of ligand-activated channels.
Journal of Experimental Biology, 1983
Synthetic proctolin increases the force but not the rate of heart contractions of Limulus in a ti... more Synthetic proctolin increases the force but not the rate of heart contractions of Limulus in a time- and dose-dependent manner. The threshold of this effect is 3 X 10−10M and the EDS0 is approximately 10−10M. At concentrations up to 10−7 M, proctolin has no effect on the rhythmic electrical activity of the isolated cardiac ganglion, and it does not change the simple and compound postsynaptic potentials recorded at the cardiac neuromuscular junction. Proctolin acts directly on the cardiac muscle fibres. Electrically stimulated myocardia show a proctolin-induced increase in contraction amplitude with the same concentration dependence as the intact heart. A compound with an apparent molecular weight of 700–800 occurs in the Limulus nervous system, particularly in the cardiac ganglion. This compound resembles proctolin in being heat-stable, resistant to trypsin and chymotrypsin cleavage, and losing activity in a time-dependent manner in response to treatment with leucine aminopeptidase ...
Journal of Experimental Biology, 1992
Mechanically isolated neuronal somata from the thoracic ganglia of the locust Locusta migratoria ... more Mechanically isolated neuronal somata from the thoracic ganglia of the locust Locusta migratoria remain electrophysiologically viable under current-or voltage-clamp in vitro for many hours. Nicotine and muscarine evoke different responses when pressure-microapplied to these somata. The response to acetylcholine is mainly nicotinic but contains a small muscarinic component. The nicotinic (AChl) response is a rapid depolarisation accompanied by a decrease in membrane resistance. In voltage-clamped somata, the current mediating the AChl response is inward over the membrane potential range −30 to − 110 mV, decreasing with depolarisation and with a projected reversal potential of about +20 mV. The muscarinic (ACh2) response is a slow depolarisation accompanied by a decrease in membrane resistance. In voltage-clamped somata, the current mediating the ACh2 response is inward, decreasing to zero at potentials of −80 to −90 mV. The AChl response is evoked by nicotine, anabasine, tetramethyla...
Journal of Experimental Biology, 1977
A relaxation oscillator, feed-back model for the circadian clock in the eye of Aplysia is propose... more A relaxation oscillator, feed-back model for the circadian clock in the eye of Aplysia is proposed to account for the experimental findings described earlier. Further data on the effects of light pulses and temperature pulses are reported here to test the hypothesis that light and temperature perturb the clock oscillation at different points in the feed-back loop. The rising phase of the CAP frequency rhythm is postulated to be due to an energy-requiring, synthesis process, and the falling phase to a passive, diffusional process. Synthesis produces a substance, C, which controls CAP frequency, and the level of which oscillates about a reference level, R. The synthesis phase of the oscillation is suggested to be temperature compensated from about 9 °C to at least 22.5 °C. Cooling the eye to 6 °C for long periods therefore inhibits synthesis so that the clock eventually stops at its lowest phase point. 12 h cold pulses of 4 °C applied during the rising phase of the rhythm (i.e. during...
Nature, 1999
The neurons were visualized by phase-contrast microscopy with an inverted microscope (Nikon Diaph... more The neurons were visualized by phase-contrast microscopy with an inverted microscope (Nikon Diaphot) while recording (in voltage clamp unless otherwise stated (V c ¼ Ϫ 70 mV) and stimulation (1 ms, +100 mV step depolarization in voltage clamp) was performed using patch-clamp amplifiers (Axopatch 200B, Axon) interfaced with a PC. Signals filtered at 5 kHz using amplifier circuitry were sampled at 10 kHz and analysed using Axoscope software (Axon). Series resistance (10-30 MQ) was always compensated at 80% (lag 100 s). For assaying synaptic connectivity, each neuron was stimulated at a low frequency (0.03-0.06 Hz), and the responses from the other neurons as well as autaptic responses in the stimulated neuron itself were recorded (see Fig. 4). Monosynaptic currents had onset latencies Ͻ4 ms (ref. 29). Polysynaptic currents had onset latencies Ն6 ms and often exhibited multiple components, with frequent failure for some PSC components to occur during the test stimulation. P of an identified component is measured based on its response to 50-100 consecutive test stimuli (0.05 Hz unless otherwise indicated). For polysynaptic pathways in these cultures, each additional synapse usually introduced a delay of 4-10 ms (2-5 ms for synaptic delay and action-potential conduction and 2-5 ms delay for the initiation of an action potential). Neurons in these cultures were either glutamate-mediating or GABA-mediating in nature and could be identified based on the time course, reversal potential and pharmacology of their evoked synaptic currents (EPSCs and IPSCs, respectively) 13,29. EPSCs were blocked by 10 M 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, RBI) whereas IPSCs were blocked by 10 M bicuculline (RBI). The IPSCs had distinctly longer decay time and more negative reversal potentials (around −50 mV) than EPSCs. In a typical culture, we estimated that less than 20% of the neurons were GABA-mediated.
Nature Neuroscience, 1998
Most fast inhibitory neurotransmission in the brain is mediated by GABA A receptors, which are ma... more Most fast inhibitory neurotransmission in the brain is mediated by GABA A receptors, which are mainly postsynaptic and consist of diverse α and β subunits together with the γ2 subunit. Although the γ2 subunit is not necessary for receptor assembly and translocation to the cell surface, we show here that it is required for clustering of major postsynaptic GABA A receptor subtypes. Loss of GABA A receptor clusters in mice deficient in the γ2 subunit, and in cultured cortical neurons from these mice, is paralleled by loss of the synaptic clustering molecule gephyrin and synaptic GABAergic function. Conversely, inhibiting gephyrin expression causes loss of GABA A receptor clusters. The γ2 subunit and gephyrin are thus interdependent components of the same synaptic complex that is critical for postsynaptic clustering of abundant subtypes of GABA A receptors in vivo.
The Journal of Physiology, 2000
GABA is the principal inhibitory neurotransmitter in the CNS, acting on ionotropic GABAA and meta... more GABA is the principal inhibitory neurotransmitter in the CNS, acting on ionotropic GABAA and metabotropic GABAB receptors. GABAA receptors are heteropentamers assembled from a repertoire of at least 19 related subunits (6 á, 3 â, 3 ã, 1 ä, 1 å, 1 ð, 3 ñ, 1 È). Many neurones express multiple subunits that give rise to a heterogeneous population of receptor subtypes. Different GABAA receptor subtypes are expressed during development, show varied regional and subcellular distribution and might contribute to short-or long-term adaptive changes in CNS function (Fritschy &
Proceedings of the National Academy of Sciences, 1999
Synaptic localization of γ-aminobutyric acid type A (GABA A ) receptors is a prerequisite for syn... more Synaptic localization of γ-aminobutyric acid type A (GABA A ) receptors is a prerequisite for synaptic inhibitory function, but the mechanism by which different receptor subtypes are localized to postsynaptic sites is poorly understood. The γ2 subunit and the postsynaptic clustering protein gephyrin are required for synaptic localization and function of major GABA A receptor subtypes. We now show that transgenic overexpression of the γ3 subunit in γ2 subunit-deficient mice restores benzodiazepine binding sites, benzodiazepine-modulated whole cell currents, and postsynaptic miniature currents, suggesting the formation of functional, postsynaptic receptors. Moreover, the γ3 subunit can substitute for γ2 in the formation of GABA A receptors that are synaptically clustered and colocalized with gephyrin in vivo . These clusters were formed even in brain regions devoid of endogenous γ3 subunit, indicating that the factors present for clustering of γ2 subunit-containing receptors are suffi...
Proceedings of the National Academy of Sciences, 1995
Vigilance, anxiety, epileptic activity, and muscle tone can be modulated by drugs acting at the b... more Vigilance, anxiety, epileptic activity, and muscle tone can be modulated by drugs acting at the benzodiazepine (BZ) site of gamma-aminobutyric acid type A (GABAA) receptors. In vivo, BZ sites are potential targets for endogenous ligands regulating the corresponding central nervous system states. To assess the physiological relevance of BZ sites, mice were generated containing GABAA receptors devoid of BZ sites. Following targeted disruption of the gamma 2 subunit gene, 94% of the BZ sites were absent in brain of neonatal mice, while the number of GABA sites was only slightly reduced. Except for the gamma 2 subunit, the level of expression and the regional and cellular distribution of the major GABAA receptor subunits were unaltered. The single channel main conductance level and the Hill coefficient were reduced to values consistent with recombinant GABAA receptors composed of alpha and beta subunits. The GABA response was potentiated by pentobarbital but not by flunitrazepam. Diazep...
Proceedings of the National Academy of Sciences, 1980
Addition of serotonin to the medium bathing an Aplysia abdominal ganglion causes a change in the ... more Addition of serotonin to the medium bathing an Aplysia abdominal ganglion causes a change in the endogenous bursting activity of the identified neuron R15. At serotonin concentrations in the micromolar range, the predominant effect is an increase in depth and duration of the interburst hyperpolarization and consequent decrease in burst rate. At higher concentrations (10 microM) serototin can inhibit bursting completely. We have shown previously that these changes can be mimicked by bath application or intracellular injection of several cyclic AMP analogs substituted at the 8 position. Voltage clamp analysis indicates that serotonin and cyclic AMP analogs both cause an increase in membrane slope conductance in R15, with reversal potentials for the responses between -75 and -80 mV, close to the K+ equilibrium potential. When the K+ concentration in the bathing medium is changed, the reversal potentials change in a manner suggesting that serotonin and cyclic AMP analogs on K+ conductan...
FEBS Letters, 1998
Amino acids in the K K-and Q Q-subunits contribute to the benzodiazepine binding site of GABA e-r... more Amino acids in the K K-and Q Q-subunits contribute to the benzodiazepine binding site of GABA e-receptors. We show that the mutation of a conserved histidine residue in the N-terminal extracellular segment (K K1 rIHI , K K2 rIHI , K K3 rIPT , and K K5 rIHS) results not only in diazepam-insensitivity of the respective K KxL L2,3Q Q2-receptors but also in an increased potentiation of the GABA-induced currents by the partial agonist bretazenil. Furthermore, Ro 15-4513, an inverse agonist at wildtype receptors, acts as an agonist at all mutant receptors. This conserved molecular switch can be exploited to identify the pharmacological significance of specific GABA e-receptor subtypes in vivo.
Nature, 2000
The quality of Fig. 2 was unsatisfactory as published. The ®gure is reproduced again here. M α1 α... more The quality of Fig. 2 was unsatisfactory as published. The ®gure is reproduced again here. M α1 α2 α3 Wild-type α1(H101R) γ2 β2/3 α5 Wild-type α1(H101R) a Wild-type α1(H101R) 28 ± 15 Diazepam > 10,000 Clonazepam 3 ± 2 > 10,000 Zolpidem 13 ± 5 > 10,000 Displacement of [ 3 H]Ro15-4513, K i (nM) c Wild-type α1(H101R) α1 b α1(H101R) Wild-type d e Wild-type 3 µM GABA+Dz +Ro15 3 µM GABA α1(H101R) 3 µM GABA+Dz +Ro15 3 µM GABA 500 pA 2 s [ 3 H]Ro15-4513 [ 3 H]Ro15-4513, K i (nM) + diazepam
Journal of Experimental Biology, 1984
Dopamine, a cardioexcitor in decapod crustaceans, increased the frequency and/or duration of burs... more Dopamine, a cardioexcitor in decapod crustaceans, increased the frequency and/or duration of bursts of action potentials in the semi-isolated cardiac ganglia of two species of crabs. The number of motoneurone action potentials in each burst was increased, which in the intact heart would increase the force and amplitude of heart contraction. The effects were concentration-dependent, with a threshold concentration of 10−8M or lower when dopamine was applied by continuous perfusion. At 5 × 10−6M, dopamine increased burst frequency by 200%. The main site of dopamine action was the group of four posterior small interneurones which normally function as the pacemaker for the cardiac ganglion system. Effects on the five large motoneurones occurred at higher concentrations. This regional difference in sensitivity was demonstrated by selective applications of dopamine to different parts of the cardiac ganglion and by the use of preparations in which the two ends of the ganglion had been funct...
Journal of Experimental Biology, 1989
The isolated, intact heart of the marine arachnid Limulus polyphemus continues to beat in vitro f... more The isolated, intact heart of the marine arachnid Limulus polyphemus continues to beat in vitro for many hours. Application of γ-aminobutyric acid (GABA) decreased the heart beat frequency with a threshold of 3×10−7moll−1 and an EC50 of 2·0±0·6×10−5moll−1 (mean±S.D., N = 8). At 10−4moll−1 and above the heart beat was completely and reversibly inhibited. The agonist potency profile of the Limulus heart chronotropic GABA receptor was very similar to that of the vertebrate GABAA receptor: muscimol > ZAPA>GABA≈TACA>isoguvacine>THIP>3-aminopropane sulphonic acid> imidazole-4-acetic acid ≈β-guanidino proprionic acid ≈5-aminovalerate. In contrast, the antagonist profile differed dramatically: bicuculline, pitrazepin and SR 95103, as well as the channel blocker picrotoxin, were without effect at concentrations up to 10−4moll−1. The benzodiazepines clorazepate, flunitrazepam, flurazepam and diazepam, as well as the barbiturate sodium pentobarbital, were without effect on th...
Journal of Experimental Biology, 1980
The five large and four small neurones in the cardiac ganglion of the crab, Portunus, are electro... more The five large and four small neurones in the cardiac ganglion of the crab, Portunus, are electrotonically coupled and behave as a single relaxation oscillator, exhibiting periodic bursting activity in vitro. Recorded from the large neurone somata, this activity consists of 200–400 ms slow depolarizations called ‘driver potentials’ (Tazaki & Cooke, 1979a), accompanied by attenuated action potentials and EPSP’s from small neurone input. There is a strong positive correlation between the duration of the driver potential and the duration of the following interburst interval in the spontaneously active ganglion. This correlation is preserved during prolonged depolarization and hyperpolarization. When a driver potential is prematurely terminated by an injected current pulse, the following interburst interval is shortened in direct proportion to the decrease in driver potential duration. When a driver potential or a burst of high-frequency action potential activity is evoked by a depolari...
Journal of Experimental Biology, 1992
1. Three different responses were evoked by pressure micro-application of serotonin onto freshly ... more 1. Three different responses were evoked by pressure micro-application of serotonin onto freshly dissociated, current- and voltage-clamped neuronal somata from the thoracic ganglia of the locust Locusta migratoria. 2. In some neurones, an inward current, I(5HT)K, resulting from a decrease in potassium conductance, with slow kinetics and maximum activation at membrane potentials of −60 to - 70 mV, was evoked by serotonin and by the 5-HT3 agonist 2-methyl serotonin. This current was completely abolished by either 10 mmoll−1 caesium or 5 mmoll−1 rubidium and partially blocked by 50 mmoll−1 tetraethylammonium or 5 mmoll−1 4-aminopyridine. The response was antagonised by the 5-HT2-specific compounds, ketanserin and ritanserin. 3. In other somata, serotonin, 2-methyl serotonin and the 5-HT3 antagonist ICS205 930 evoked a second current, I(5HT)Na, which was due to an increase in sodium permeability and had slow kinetics similar to that of I(5HT)K. This current was inward over the membrane ...
Trends in Neurosciences, 1984
The organization into bursts of action potential firing is a characteristic of the neurons of man... more The organization into bursts of action potential firing is a characteristic of the neurons of many groups of animals. In the crustacean cardiac and stomatogastric ganglia, burst generation and perhaps also endogenous rhythmiciry result from the production of slow, &+-dependent, non-propagated depolarizations known as driver or plateau potentials. At least some neuromodulatory inputs to these ganglia have their effects by acting on these potentials, and driver potential formation appears to be an essential aspect of non-impulse-mediated synaptic coordination in these ganglia.
Proceedings of the National Academy of Sciences, 1983
Previous work has shown that serotonin causes an increase in K+ conductance in the identified Apl... more Previous work has shown that serotonin causes an increase in K+ conductance in the identified Aplysia neuron R15. This response is mediated by cAMP-dependent protein phosphorylation. The results presented here show that the K+ channel modulated by serotonin is an anomalous or inward rectifier (designated IR) that is present in R15 together with the three other distinct K+ channels previously described for this cell. Several lines of evidence indicate that this inward rectifier is partially activated in the resting cell and is further activated by serotonin. Voltage clamp analysis of resting and serotonin-evoked membrane currents at various external K+ concentrations shows that both currents have reversal potentials close to the potassium equilibrium potential, exhibit similar dependences in magnitude on external K+ concentration, and display marked anomalous rectification. The effects of particular monovalent and divalent cations are also similar on the resting and serotonin-evoked ...
European Journal of Pharmacology, 2001
The functional pharmacology of receptors composed of the chicken brain GABA receptor g4 subunit a... more The functional pharmacology of receptors composed of the chicken brain GABA receptor g4 subunit and the mammalian GABA A A receptor a 3 and b2 subunits was studied by heterologous expression in Xenopus laeÕis oocytes using the two electrode voltage-clamp technique. GABA-evoked currents had an EC of 180 " 30 mM. Responses were blocked by the competitive and non-competitive 50 GABA receptor antagonists, bicuculline methochloride and picrotoxin. Sodium pentobarbital reversibly potentiated the current A 2q Ž. several-fold, and Zn ions blocked the current with high potency IC s 20 mM. GABA-evoked currents were potentiated by the 50 benzodiazepine site full agonists flunitrazepam and triazolam and less by the partial agonists abecarnil and bretazenil. The inverse Ž. Ž. agonists methyl-b-carboline-3-carboxylate b-CCM and methyl 6,7-dimethoxy-4-ethyl-b-carboline-3-carboxylate DMCM reduced the current. However, the imidazobenzodiazepine Ro 15-4513, which acts as an inverse agonist at mammalian a xb y g2 GABA receptors A
Brain Research, 1988
The isolated somata of neurons from the thoracic ganglia of the locust, Locusta migratoria, respo... more The isolated somata of neurons from the thoracic ganglia of the locust, Locusta migratoria, respond to pressure microapplication of y-aminobutyric acid (GABA) and acetylcholine. The acetylcholine receptors fall into two groups, ACh t (activated by nicotine) and ACh 2 (activated by muscarine). The GABA receptor and the ACh I receptor differ in pharmacology from the known vertebrate receptors. The GABA receptor is insensitive to bicuculline and its salts up to a concentration of 10-4 M. In contrast, bicuculline is a moderately potent, at least partially competitive antagonist of the ACh~ receptor-mediated response in the thoracic neuronal somata. These observations suggest that classical diagnostic compounds such as bicuculline may show greater cross-reactivity than hitherto suspected among the members of the superfamily of ligand-activated channels.
Journal of Experimental Biology, 1983
Synthetic proctolin increases the force but not the rate of heart contractions of Limulus in a ti... more Synthetic proctolin increases the force but not the rate of heart contractions of Limulus in a time- and dose-dependent manner. The threshold of this effect is 3 X 10−10M and the EDS0 is approximately 10−10M. At concentrations up to 10−7 M, proctolin has no effect on the rhythmic electrical activity of the isolated cardiac ganglion, and it does not change the simple and compound postsynaptic potentials recorded at the cardiac neuromuscular junction. Proctolin acts directly on the cardiac muscle fibres. Electrically stimulated myocardia show a proctolin-induced increase in contraction amplitude with the same concentration dependence as the intact heart. A compound with an apparent molecular weight of 700–800 occurs in the Limulus nervous system, particularly in the cardiac ganglion. This compound resembles proctolin in being heat-stable, resistant to trypsin and chymotrypsin cleavage, and losing activity in a time-dependent manner in response to treatment with leucine aminopeptidase ...
Journal of Experimental Biology, 1992
Mechanically isolated neuronal somata from the thoracic ganglia of the locust Locusta migratoria ... more Mechanically isolated neuronal somata from the thoracic ganglia of the locust Locusta migratoria remain electrophysiologically viable under current-or voltage-clamp in vitro for many hours. Nicotine and muscarine evoke different responses when pressure-microapplied to these somata. The response to acetylcholine is mainly nicotinic but contains a small muscarinic component. The nicotinic (AChl) response is a rapid depolarisation accompanied by a decrease in membrane resistance. In voltage-clamped somata, the current mediating the AChl response is inward over the membrane potential range −30 to − 110 mV, decreasing with depolarisation and with a projected reversal potential of about +20 mV. The muscarinic (ACh2) response is a slow depolarisation accompanied by a decrease in membrane resistance. In voltage-clamped somata, the current mediating the ACh2 response is inward, decreasing to zero at potentials of −80 to −90 mV. The AChl response is evoked by nicotine, anabasine, tetramethyla...
Journal of Experimental Biology, 1977
A relaxation oscillator, feed-back model for the circadian clock in the eye of Aplysia is propose... more A relaxation oscillator, feed-back model for the circadian clock in the eye of Aplysia is proposed to account for the experimental findings described earlier. Further data on the effects of light pulses and temperature pulses are reported here to test the hypothesis that light and temperature perturb the clock oscillation at different points in the feed-back loop. The rising phase of the CAP frequency rhythm is postulated to be due to an energy-requiring, synthesis process, and the falling phase to a passive, diffusional process. Synthesis produces a substance, C, which controls CAP frequency, and the level of which oscillates about a reference level, R. The synthesis phase of the oscillation is suggested to be temperature compensated from about 9 °C to at least 22.5 °C. Cooling the eye to 6 °C for long periods therefore inhibits synthesis so that the clock eventually stops at its lowest phase point. 12 h cold pulses of 4 °C applied during the rising phase of the rhythm (i.e. during...
Nature, 1999
The neurons were visualized by phase-contrast microscopy with an inverted microscope (Nikon Diaph... more The neurons were visualized by phase-contrast microscopy with an inverted microscope (Nikon Diaphot) while recording (in voltage clamp unless otherwise stated (V c ¼ Ϫ 70 mV) and stimulation (1 ms, +100 mV step depolarization in voltage clamp) was performed using patch-clamp amplifiers (Axopatch 200B, Axon) interfaced with a PC. Signals filtered at 5 kHz using amplifier circuitry were sampled at 10 kHz and analysed using Axoscope software (Axon). Series resistance (10-30 MQ) was always compensated at 80% (lag 100 s). For assaying synaptic connectivity, each neuron was stimulated at a low frequency (0.03-0.06 Hz), and the responses from the other neurons as well as autaptic responses in the stimulated neuron itself were recorded (see Fig. 4). Monosynaptic currents had onset latencies Ͻ4 ms (ref. 29). Polysynaptic currents had onset latencies Ն6 ms and often exhibited multiple components, with frequent failure for some PSC components to occur during the test stimulation. P of an identified component is measured based on its response to 50-100 consecutive test stimuli (0.05 Hz unless otherwise indicated). For polysynaptic pathways in these cultures, each additional synapse usually introduced a delay of 4-10 ms (2-5 ms for synaptic delay and action-potential conduction and 2-5 ms delay for the initiation of an action potential). Neurons in these cultures were either glutamate-mediating or GABA-mediating in nature and could be identified based on the time course, reversal potential and pharmacology of their evoked synaptic currents (EPSCs and IPSCs, respectively) 13,29. EPSCs were blocked by 10 M 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, RBI) whereas IPSCs were blocked by 10 M bicuculline (RBI). The IPSCs had distinctly longer decay time and more negative reversal potentials (around −50 mV) than EPSCs. In a typical culture, we estimated that less than 20% of the neurons were GABA-mediated.
Nature Neuroscience, 1998
Most fast inhibitory neurotransmission in the brain is mediated by GABA A receptors, which are ma... more Most fast inhibitory neurotransmission in the brain is mediated by GABA A receptors, which are mainly postsynaptic and consist of diverse α and β subunits together with the γ2 subunit. Although the γ2 subunit is not necessary for receptor assembly and translocation to the cell surface, we show here that it is required for clustering of major postsynaptic GABA A receptor subtypes. Loss of GABA A receptor clusters in mice deficient in the γ2 subunit, and in cultured cortical neurons from these mice, is paralleled by loss of the synaptic clustering molecule gephyrin and synaptic GABAergic function. Conversely, inhibiting gephyrin expression causes loss of GABA A receptor clusters. The γ2 subunit and gephyrin are thus interdependent components of the same synaptic complex that is critical for postsynaptic clustering of abundant subtypes of GABA A receptors in vivo.
The Journal of Physiology, 2000
GABA is the principal inhibitory neurotransmitter in the CNS, acting on ionotropic GABAA and meta... more GABA is the principal inhibitory neurotransmitter in the CNS, acting on ionotropic GABAA and metabotropic GABAB receptors. GABAA receptors are heteropentamers assembled from a repertoire of at least 19 related subunits (6 á, 3 â, 3 ã, 1 ä, 1 å, 1 ð, 3 ñ, 1 È). Many neurones express multiple subunits that give rise to a heterogeneous population of receptor subtypes. Different GABAA receptor subtypes are expressed during development, show varied regional and subcellular distribution and might contribute to short-or long-term adaptive changes in CNS function (Fritschy &
Proceedings of the National Academy of Sciences, 1999
Synaptic localization of γ-aminobutyric acid type A (GABA A ) receptors is a prerequisite for syn... more Synaptic localization of γ-aminobutyric acid type A (GABA A ) receptors is a prerequisite for synaptic inhibitory function, but the mechanism by which different receptor subtypes are localized to postsynaptic sites is poorly understood. The γ2 subunit and the postsynaptic clustering protein gephyrin are required for synaptic localization and function of major GABA A receptor subtypes. We now show that transgenic overexpression of the γ3 subunit in γ2 subunit-deficient mice restores benzodiazepine binding sites, benzodiazepine-modulated whole cell currents, and postsynaptic miniature currents, suggesting the formation of functional, postsynaptic receptors. Moreover, the γ3 subunit can substitute for γ2 in the formation of GABA A receptors that are synaptically clustered and colocalized with gephyrin in vivo . These clusters were formed even in brain regions devoid of endogenous γ3 subunit, indicating that the factors present for clustering of γ2 subunit-containing receptors are suffi...
Proceedings of the National Academy of Sciences, 1995
Vigilance, anxiety, epileptic activity, and muscle tone can be modulated by drugs acting at the b... more Vigilance, anxiety, epileptic activity, and muscle tone can be modulated by drugs acting at the benzodiazepine (BZ) site of gamma-aminobutyric acid type A (GABAA) receptors. In vivo, BZ sites are potential targets for endogenous ligands regulating the corresponding central nervous system states. To assess the physiological relevance of BZ sites, mice were generated containing GABAA receptors devoid of BZ sites. Following targeted disruption of the gamma 2 subunit gene, 94% of the BZ sites were absent in brain of neonatal mice, while the number of GABA sites was only slightly reduced. Except for the gamma 2 subunit, the level of expression and the regional and cellular distribution of the major GABAA receptor subunits were unaltered. The single channel main conductance level and the Hill coefficient were reduced to values consistent with recombinant GABAA receptors composed of alpha and beta subunits. The GABA response was potentiated by pentobarbital but not by flunitrazepam. Diazep...
Proceedings of the National Academy of Sciences, 1980
Addition of serotonin to the medium bathing an Aplysia abdominal ganglion causes a change in the ... more Addition of serotonin to the medium bathing an Aplysia abdominal ganglion causes a change in the endogenous bursting activity of the identified neuron R15. At serotonin concentrations in the micromolar range, the predominant effect is an increase in depth and duration of the interburst hyperpolarization and consequent decrease in burst rate. At higher concentrations (10 microM) serototin can inhibit bursting completely. We have shown previously that these changes can be mimicked by bath application or intracellular injection of several cyclic AMP analogs substituted at the 8 position. Voltage clamp analysis indicates that serotonin and cyclic AMP analogs both cause an increase in membrane slope conductance in R15, with reversal potentials for the responses between -75 and -80 mV, close to the K+ equilibrium potential. When the K+ concentration in the bathing medium is changed, the reversal potentials change in a manner suggesting that serotonin and cyclic AMP analogs on K+ conductan...
FEBS Letters, 1998
Amino acids in the K K-and Q Q-subunits contribute to the benzodiazepine binding site of GABA e-r... more Amino acids in the K K-and Q Q-subunits contribute to the benzodiazepine binding site of GABA e-receptors. We show that the mutation of a conserved histidine residue in the N-terminal extracellular segment (K K1 rIHI , K K2 rIHI , K K3 rIPT , and K K5 rIHS) results not only in diazepam-insensitivity of the respective K KxL L2,3Q Q2-receptors but also in an increased potentiation of the GABA-induced currents by the partial agonist bretazenil. Furthermore, Ro 15-4513, an inverse agonist at wildtype receptors, acts as an agonist at all mutant receptors. This conserved molecular switch can be exploited to identify the pharmacological significance of specific GABA e-receptor subtypes in vivo.