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S G Cull-Candy

Publications and source records attributed to S G Cull-Candy.

At least 55 records · Page 3Linked to original sources
PubMed1989

On the multiple-conductance single channels activated by excitatory amino acids in large cerebellar neurones of the rat.

1. Single-channel currents evoked by excitatory amino acids have been examined in outside-out patches from large cerebellar neurones (including Purkinje cells) in tissue culture. L-Glutamate (3-10 microM), L-aspartate (3-10 microM), NMDA (N-methyl-D-aspartate, 10-50 microM), ibotenate (50 microM), quisqualate (3-50 microM), and kainate (3-50 microM) all produced single-channel currents with multiple amplitudes. 2. Single-channel currents recorded over a range of patch potentials had a mean interpolated reversal potential of -3.8 +/- 0.5 mV. The directly resolvable multiple conductance levels could be classified into five main groups, with mean values (averaged for all agonists) of: 47.9 +/- 0.7, 38.5 +/- 0.8, 27.8 +/- 1.4, 18.2 +/- 0.5 and 8.3 +/- 0.6 pS. 3. From the relative areas under current amplitude histograms it was estimated that the percentage of openings with conductances greater than 30 pS was about 83% with NMDA, 79% with glutamate and 78% with aspartate. In some patches, the majority of greater than 30 pS events evoked by these agonists were to the maximum conductance of 48 pS, whereas in other patches there were more 38 pS openings than 48 pS openings. Only 27% of quisqualate openings, and about 10% of kainate openings, were greater than 30 pS. 4. Of the small amplitude (less than 20 pS) events, 93% of quisqualate openings were to the 8 pS level whereas approximately 87% of less than 20 pS currents produced by NMDA, glutamate and aspartate were to the 18 pS level (the remainder being 8 pS). Direct transitions could occur between certain levels (including events above and below 30 pS) suggesting that these are sublevels of multiple-conductance channels. The most frequently occurring transitions were between the 48 and 38 pS levels, and the 38 and 18 pS levels. 5. Channel openings occurred in bursts, within which individual openings were separated either by brief closures (gaps), or by direct transitions between the multiple conductance levels. The briefest of these gaps (less than 200-400 microseconds) could represent a mixture of transitions to lower conductance levels as well as partially resolved complete shuttings. The mean duration of the longer gaps within bursts, thought to represent complete but partially resolved shuttings was 1.05 +/- 0.25 ms (pooled for all agonists). 6. Burst-length distributions could be fitted with the sum of three exponentials. The briefest component may have arisen from brief single openings. The two slower components probably reflect the existence of two kinetically distinct open states.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1988

On the kinetics of large-conductance glutamate-receptor ion channels in rat cerebellar granule neurons.

Ion channels activated by glutamate, aspartate, and N-methyl-D-aspartate (NMDA) have been investigated in outside-out patches from cultured cerebellar granule neurons of the rat. Openings of these channels occur in bursts, within which the individual openings are separated by brief shuttings or gaps. The shut-time distributions obtained with each agonist were fitted with four exponential components. The briefest two components were considered as 'gaps within bursts'. Their mean time-constants were: glutamate, 58.0 microseconds and 592 microseconds; aspartate, 31.3 microseconds and 644 microseconds; NMDA, 40.5 microseconds and 903 microseconds. Distributions of burst durations were fitted with three exponential components. The mean time-constants obtained for the longest two components were: glutamate, 1.33 ms and 10.5 ms; aspartate, 2.15 ms and 10.3 ms; NMDA, 2.42 ms and 10.5 ms. Evidence is given that these two components of burst duration reflect the gating kinetics of 50 pS openings and not the fact that each agonist produces openings to more than one conductance level. Not only do openings occur in bursts, but these bursts were observed to occur in clusters, which can be hundreds of milliseconds long. We discuss the relation between the kinetics of single-channel openings observed in patches and the spectral components detected in whole-cell current noise.

Animals↗
PubMed1988

Noise and single channels activated by excitatory amino acids in rat cerebellar granule neurones.

1. Glutamate-receptor ion channels in rat cerebellar granule cells maintained in explant cultures have been investigated with patch-clamp methods. Properties of these channels were determined from noise analysis of whole-cell currents and from noise and single-channel currents recorded in outside-out membrane patches. 2. Glutamate (10-20 microM) evoked two types of response. Some granule cells gave small inward currents accompanied by clear increases in current noise ('large noise' responses), whereas other cells gave larger inward currents and small noise increases ('small noise' responses). 3. A mean single-channel conductance (gamma) of 46.6 pS was estimated for glutamate from four 'large noise' cells. A mean gamma value of 8.4 pS was estimated for seven other 'large noise' cells. The results suggest that in these latter cells glutamate activated both large (approximately equal to 50 pS) and small conductance (approximately equal to 140 fS) channels. 4. Applications of aspartate (10-30 microM) or N-methyl-D-aspartate (NMDA, 10-30 microM) produced small inward currents and large increases in noise; gamma noise = 48.5 pS (aspartate) and 46.7 pS (NMDA). 5. Large single-channel currents were evoked by glutamate, aspartate and NMDA in outside-out patches. The mean conductance values obtained for the largest amplitude openings were: gamma(glutamate) = 49.5 pS, gamma(aspartate) = 51.5 pS, and gamma(NMDA) = 53.0 pS. For each agonist, these 50 pS openings comprised 75-85% of the completely resolved currents in each patch. Openings to 40 and 30 pS conductance levels accounted for 10-15% and 3-7% of the total, and the presence of apparently direct transitions between these levels and the 50 pS level suggests they are sublevels of the same multi-conductance channels. 6. A mean channel conductance of 22.9 pS was estimated from noise evoked by quisqualate (10-30 microM). Single-channel currents were examined in four patches. In two, quisqualate evoked predominantly small currents of two amplitudes, gamma = 8.4 pS and 16.5 pS; some 50 pS openings were also present. In the other two patches, most openings were 50 pS events. 7. Granule cells gave inward currents to kainate (10-30 microM), and a mean conductance of 3.1 pS was estimated from kainate noise. In patches in which aspartate or NMDA produced mainly 50 pS openings, more than 74% of the single-channel currents evoked by kainate were of smaller amplitude, with mean conductances of gamma = 8.1 and 15.1 pS.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1988

Acetylcholine receptor channels and their block by clonidine in cultured bovine chromaffin cells.

1. Acetylcholine (10-50 microM) applied to bovine chromaffin cells under whole-cell voltage clamp produced an inward current and an increase in the noise level of the current trace. At concentrations greater than or equal to 20 microM the ACh current desensitized with time. Spectral analysis of the ACh-induced increase in current noise showed that it could be fitted by a single Lorentzian component with a time constant, tau noise = 11 +/- 0.9 ms (Vm = -80 mV). 2. Clonidine (2-30 microM) markedly reduced the size of the ACh-induced whole-cell current, and altered the shape of the noise spectrum. In the presence of clonidine, ACh noise spectra were fitted by two Lorentzian components with time constants which varied with clonidine concentration. The single-channel conductance from noise (gamma = 24 pS) was unaltered by clonidine. 3. The reduction in the size of the whole-cell ACh current, produced by clonidine, was not mimicked by adrenaline (at concentrations up to 60 microM). GABA-induced whole-cell currents and spectra of GABA noise were also unchanged suggesting that the effect of clonidine was specific for the ACh receptor channel. 4. When applied intracellularly (from the patch pipette) clonidine had no apparent influence on the whole-cell ACh current. Furthermore, when clonidine was perfused over the cell surface at a concentration sufficient to block the whole-cell ACh response, single ACh channels could still be recorded under the patch-pipette tip (i.e. in cell-attached patches, not exposed to clonidine). 5. Clonidine block showed little voltage dependence; the peak ACh current remained linearly dependent on clamp potential. In addition, the fast and slow time constants derived from ACh noise spectra in the presence of clonidine did not show the sort of dependence on antagonist concentration expected for simple channel block; this suggests that clonidine has a complex blocking action. 6. Single ACh channels recorded in outside-out patches had a conductance of gamma = 39 +/- 0.7 pS, although a subpopulation of smaller and larger events also occurred in some patches. The mean single-channel conductance in outside-out patches was unaltered by clonidine. 7. Under normal conditions the kinetics of single ACh channels were more complex than suggested from noise analysis. Burst length distributions could be described by two exponential components with fast and slow time constants of tau f = 0.69 +/- 0.17 and tau s = 9.51 +/- 0.84 ms.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcholine↗
PubMed1987

Single-channel and whole-cell currents evoked by acetylcholine in dissociated sympathetic neurons of the rat.

Single acetylcholine-activated channels have been recorded from neurons dissociated from the sympathetic chain of 17-21 day old rats. The mean single channel conductance is 35 pS in normal medium containing 1 mM calcium, and 51 pS in the absence of calcium. The measured current amplitudes are about five times more variable than at the frog endplate, at least in part because the current, while the channel is open, is much noisier than when it is shut. Single activations of the receptor by acetylcholine (ACh) produce a burst of openings; the distribution of the burst length has two components, the longer of which is of primary importance in synaptic transmission. Whole-cell currents, in response to ACh (up to 30 microM), show strong inward rectification with no outward current being detectable. This phenomenon is similar whether the intracellular ion is sodium or cesium, whether or not divalent cations are present, and whether or not atropine is present. Nevertheless, outward single-channel currents (of normal conductance) are detectable in isolated outside-out patches.

Acetylcholine↗
PubMed1987

Glutamate and aspartate activated channels and inhibitory synaptic currents in large cerebellar neurons grown in culture.

Patch-clamp methods were used to examine large (greater than 30 microns) cerebellar neurons of the rat, maintained in cell culture. Cells possessed voltage-activated transient inward Na+ currents which were sensitive to tetrodotoxin. Spontaneous synaptic currents, present in whole-cell recordings, were abolished by bicuculline and picrotoxin and were carried by Cl-. Cells produced inward currents in response to the transmitter candidates glutamate and aspartate and also to the glutamate agonists kainate, quisqualate, N-methyl-D-aspartate and ibotenate. Analysis of glutamate and aspartate-current noise has been used to derive characteristics of the excitatory channels. Single channel currents have also been observed directly in whole-cell and outside-out patches. Both glutamate and aspartate are able to activate channels which were blocked by Mg2+ and had a maximum conductance of 50 pS.

Animals↗
PubMed1986

Ion channels activated by acetylcholine and gamma-aminobutyric acid in freshly dissociated sympathetic neurones of the rat.

Transmitter-activated channels in freshly isolated neurones from sympathetic ganglia of young rats have been examined using the patch-clamp technique. Acetylcholine (ACh), gamma-aminobutyric acid (GABA) and (in about one third of cells) glycine produced an inward current and an increase in current noise when perfused onto voltage-clamped neurones. The ACh noise was fitted with a single Lorentzian spectrum with a time constant of 13.1 ms at -70 mV. Outside-out membrane patches allowed high-resolution measurements of single ACh- and GABA-activated channels. The ACh-channels had a conductance of 30 pS.

Acetylcholine↗
PubMed1986

Miniature and evoked inhibitory junctional currents and gamma-aminobutyric acid-activated current noise in locust muscle fibres.

gamma-Aminobutyric acid (GABA) current noise and inhibitory junctional currents (i.j.c.s) have been examined to give properties of the GABA receptor and its associated synaptic channel. Various procedures were used to identify muscle bundles receiving inhibitory innervation. In normal bathing medium the decay time constant of the i.j.c. was tau i.j.c. = 7.6 +/- 0.7 ms (clamp potential, Vm = -80 mV; temperature, T = 21 degrees C). Most muscle fibres were sensitive to ionophoretically applied GABA, irrespective of the presence of inhibitory innervation. GABA current noise obtained at junctional sites gave spectra which were fitted usually with a single Lorentzian component, or occasionally with the sum of two Lorentzians. The conductance of the single inhibitory channel was, gamma (GABA) = 21.6 +/- 0.9 pS (Vm = -80 mV; T = 21 degrees C). The mean 'burst length' of the openings produced by a single receptor activation was tau noise = 4.0 +/- 0.8 ms, at Vm = -80 mV. This decreased exponentially with hyperpolarization. On average tau i.j.c. exceeded tau noise although good agreement was found in some fibres. I.j.c.s were examined in greater detail after excitatory synaptic receptors had been desensitized with 10(-3) M-L-glutamate to abolish all excitatory synaptic activity. Their decay time constant was tau i.j.c. = 7.2 +/- 0.4 ms, and their rise time was 3.3 +/- 0.12 ms, at Vm = -80 mV. An e-fold decrease in tau i.j.c. resulted from a 103 +/- 7.9 mV hyperpolarization; time to peak showed a smaller dependence on Vm. The mean size of the inhibitory quantal event (i.e. response to a single transmitter packet) was estimated from fluctuations in i.j.c. amplitude. Mean quantal content of the i.j.c. was about 30 at normal levels of release. Mean amplitude of the directly measured miniature i.j.c. = 0.65 +/- 0.08 nA at Vm = -80 mV (V eq approximately equal to -40 mV). The amplitude of the quantal event showed a non-linear dependence on Vm. The burst length of the inhibitory channel, produced by a single receptor activation, is longer in duration (at -80 mV) and exhibits greater voltage dependence than the burst length of the excitatory glutamate-activated channel in these fibres. It is estimated that a single quantum of GABA opens about 600-1000 post-synaptic chloride channels.

Animals↗
PubMed1985

Ion channels activated by L-glutamate and GABA in cultured cerebellar neurons of the rat.

Glutamate and GABA-receptor channels were investigated in explants of rat cerebellum grown in cell culture. The patch-clamp technique was used to examine neurons under whole cell clamp and the properties of channels were derived by analysis of glutamate and GABA-evoked current noise. In addition, single channel currents activated by glutamate were recorded from isolated outside-out patches of membrane. We found evidence for at least two types of glutamate receptor-channels in cerebellar cells. Some neurons exhibited a channel of 50 pS conductance with a Lorentzian noise spectrum of 5.9 ms time constant. Single channels were readily resolved both in whole cell clamp and excised patches. Other neurons possessed low conductance channels which produced two component spectra. Estimates of the single channel conductance gave a value of about 140 fS. GABA channel noise obtained from these cells was also fitted by two component spectra which gave single channel conductance of 16 pS.

Animals↗
PubMed1984

Inhibitory synaptic currents in voltage-clamped locust muscle fibres desensitized to their excitatory transmitter.

Inhibitory junctional currents (i.j.c.s) have been examined in locust muscle fibres to give properties of GABA-channels activated by the neurally released transmitter. A nerve-muscle preparation is described which has proved suitable for voltage-clamp analysis of inhibitory transmission. I.j.c.s were recorded from fibres in which excitatory synapses had been desensitized with glutamate, to abolish excitatory junctional currents. This procedure had no apparent effect on inhibitory channel properties. The time constant of decay of the i.j.c. was 7.7 +/- 0.3 ms, slightly exceeding the time constant of the membrane noise induced by externally applied GABA. Peak i.j.c. conductance decreased with hyperpolarization. I.j.c.s showed measurable fluctuations permitting an estimate of the mean size of the quantal events composing the i.j.c. Their mean size coincided with the spontaneously occurring miniature inhibitory junctional currents that could be directly recorded in some fibres. The inhibitory nerve-impulse released an average of 35 transmitter packets at sites distributed along the muscle fibre length. Since each m.i.j.c. produced a current of about 0.6 nA (at Vm = -80 mV, ECl = -40 mV) the single quantum of inhibitory transmitter opens 600-1000 postsynaptic chloride channels. This is roughly three to four times the number of channels opened by the excitatory transmitter packet at glutamate synapses in the same fibres.

Animals↗
PubMed1983

Block of glutamate-activated synaptic channels by curare and gallamine.

Excitatory junctional currents (e.j.cs) and glutamate-activated currents have been examined in voltage-clamped locust muscle fibres exposed to curare or gallamine. Although these drugs have little action on channel kinetics at the resting potential, there is an increasingly pronounced effect at hyperpolarized levels. In the presence of curare (5-100 microM), fibres held at hyperpolarized potentials showed e.j.cs with an initial 'fast component' followed by a 'slow tail'. In many fibres, hyperpolarization beyond -50 mV decreased the amplitude of the peak synaptic current; the decay time constant of the fast component was decreased by hyperpolarization while the time constant of the slow component was increased. Iontophoretic application of brief pulses of glutamate also produced two-component glutamate currents in these conditions. Gallamine was considerably more effective than curare, markedly altering the decay time and amplitude of the e.j.c. and the glutamate current at 1-5 microM. Its effects appeared qualitatively similar to those of curare. The observations are consistent with the idea that curare and gallamine produce a transient block of glutamate-activated synaptic channels.

Animals↗
PubMed1982

Properties of miniature excitatory junctional currents at the locust nerve-muscle junction.

1. Miniature excitatory junctional currents (m.e.j.c.s) were examined in conditions where inward current was carried mainly by Na(+) (i.e. in normal medium, Ca(2+)-free medium and Cl(-)-free medium). M.e.j.c.s were also examined in isotonic Ca(2+) where the inward post-synaptic current was carried mainly by Ca(2+).2. In normal medium, mean m.e.j.c. amplitude = 2.34+/-0.05 nA. The decay time constant of m.e.j.c.s (excluding a small percentage with abnormal shapes) was tau(m.e.j.c.) = 2.62+/-0.11 msec (V(m) = -80 mV, T = 22 degrees C). Decay-time was not markedly changed in Ca(2+)-free or Cl(-)-free medium. tau(m.e.j.c.) approaches the life-time of glutamate activated junctional channels.3. Excitatory junctional currents, evoked by nerve impulses, decayed slightly faster than m.e.j.c.s obtained in the same fibres. Extracellularly recorded m.e.j.c.s and voltage-clamped m.e.j.c.s were similar in time course.4. tau(m.e.j.c.) decreased exponentially with membrane hyperpolarization. An e-fold change was produced by 182.+/-24.8 mV change in V(m).5. The dependence of mean m.e.j.c. amplitude on clamp potential showed a slight non-linearity at hyperpolarized levels. The equilibrium potential for transmitter action was close to 0 mV in normal solution as well as in Ca(2+)-free and Cl(-)-free solutions.6. The kinetics of junctional channels are altered in isotonic Ca(2+). M.e.j.c. amplitude was reduced to about one-third normal size; mean m.e.j.c. = 0.74+/-0.03 nA. The decay time becomes markedly briefer, tau(m.e.j.c.) = 1.01+/-0.08 msec, indicating a reduction in mean channel life-time (V(m) = -80 mV, T = 22 degrees C).7. A population of slow time course and composite m.e.j.c.s appear when muscle fibres are hyperpolarized in isotonic Ca(2+), thus producing a prolongation in mean tau(m.e.j.c.). This results from an influence of post-synaptic membrane potential on presynaptic transmitter release. If such m.e.j.c.s are ignored the voltage dependence of tau(m.e.j.c.) of the remaining events is abolished or even reversed indicating that voltage sensitivity of channel life-time is altered in isotonic Ca(2+). The equilibrium potential for transmitter action may be slightly more positive than normal.8. We estimate that a single packet of neurally released transmitter normally opens, on average, 250 ion channels at these junctions.

Animals↗
PubMed1982

Denervation changes in normal and myasthenia gravis human muscle fibres during organ culture.

1. Human intercostal nerve-muscle obtained from normal and myasthenia gravis affected patients has been organ cultured for up to 5 weeks at 23 degrees C. In addition normal nerve-muscle has been cultured for up to 2 weeks at 36 degrees C. Muscle fibres had normal resting and overshooting action potentials. Input resistances dropped markedly after 21 days at 23 degrees C or 10 days at 36 degrees C.2. Muscle fibre action potentials became partially resistant to tetrodotoxin (10(-7) g/ml.) after culturing for 8 days at 36 degrees C.3. Extrajunctional acetylcholine (ACh) sensitivity was compared in fresh and cultured muscle. Fresh normal fibres possessed extrajunctional ACh sensitivity covering several hundred micrometres around the end-plate and at the muscle-tendon junction. Myasthenia gravis affected fibres had reduced extrajunctional sensitivity at the end-plate and no detectable ACh sensitivity near the tendon.4. Less than one third of normal muscle bundles showed an increased area of ACh sensitivity after several days in organ culture. Under the same conditions myasthenic muscle bundles did not show an increase in extrajunctional ACh sensitivity,5. M.e.p.p.s were present for 2-4 days in normal fibres cultured at 36 degrees C. In normal and myasthenic muscles cultured at 23 degrees C m.e.p.p.s disappeared after 6-8 days and re-appeared in some fibres (50%) after approximately two weeks in organ culture. These m.e.p.p.s were abolished by curare and increased in frequency by hypotonic solution suggesting they are due to the release of ACh-packets from Schwann cells.6. Electron microscopic examination of cultured human muscle indicates that the disappearance of m.e.p.p.s corresponds with degeneration of nerve terminals.7. In muscle bundles shown to possess m.e.p.p.s after 13-14 days, the synaptic gutter, which had been vacated by the nerve terminal, was usually occupied by a Schwann cell or projections of Schwann cell cytoplasm. This indicates that the Schwann cell at denervated human end-plates may be capable of releasing packets of ACh.8. It is concluded that the organ culture system described here is suitable for studying normal and diseased human muscle fibres. Using this system we find that the denervation changes which follow nerve transection appear to be similar in most respects at normal and myasthenic end-plates.

Action Potentials↗
PubMed1982

Properties of junctional and extrajunctional acetylcholine-receptor channels in organ cultured human muscle fibres.

1. Current noise, obtained during steady ionophoretic application of acetylcholine (ACh) to voltage-clamped human fibres has been analysed to derive properties of end-plate channels and also extrajunctional ACh-activated channels which are present at the muscle-tendon junction of normal fibres. In addition, ACh-receptor channels present at the end-plate and tendon region in organ cultured muscles have been compared with those in fresh muscles.2. Extrajunctional channels in the tendon region of fresh fibres have a longer mean life-time, tau, and a smaller single channel conductance, gamma, than the junctional channels. tau was 1.71 +/-0.11 msec and gamma was 25.05 +/- 1.18 pS for junctional channels; tau was 3.16 +/- 0.33 msec and gamma was 12.76 +/- 1.29 pS for extrajunctional channels.3. Properties of channels in the end-plate and tendon region were unchanged during short-term (< 7 days) organ culture at 23 or 36 degrees C. The voltage sensitivity of the mean channel life-time was similar at junctional and extrajunctional sites.4. In muscles organ cultured for 7 days at 36 degrees C, double component noise spectra were obtained at some end-plates. The fast and slow time constants underlying the noise appeared to correspond to simultaneous activation of junctional and extrajunctional channels.5. After organ culture for 3-4 weeks at 23 degrees C the mean life-time of the end-plate channels was prolonged while their single channel conductance was unchanged, tau = 3.58 +/- 0.16 msec; gamma = 22.11 +/- 0.83 pS (V(m) = -80 mV, T = 21 degrees C).6. As end-plate channel properties were unchanged in short-term cultures it was possible to assess post-junctional sensitivity by comparing miniature end-plate current (m.e.p.c.) amplitudes in both normal and myasthenia gravis affected muscles. No evidence was found for a change in post-synaptic sensitivity during organ culture at 23 degrees C of normal or myasthenia gravis muscle fibres, which seems to rule out reversible block of receptors by anti-receptor antibody as playing an important role in myasthenia gravis.

Acetylcholine↗
PubMed1981

Junctional and extrajunctional membrane channels activated by GABA in locust muscle fibres.

Iontophoretic application of GABA to voltage-clamped locust muscle fibres has demonstrated the presence of both extrajunctional and junctional GABA receptors. Extrajunctional GABA receptors are distinct from extrajunctional glutamate receptors which also occur in these muscle fibres. Inward GABA currents are nonlinearly dependent on membrane potential. Analysis of membrane current noise produced by iontophoretic GABA application shows that for junctional and extrajunctional GABA receptors the mean channel lifetime is 3-4 ms and the single-channel conductance is approximately 22 pS at - 80 mV (T = 21 degrees C). The mean lifetime as previously demonstrated for glutamate-sensitive excitatory channels in locust muscle fibres.

Animals↗