Molecular dynamics of the sodium channel pore vary with gating: interactions between P-segment motions and inactivation.
(1/2595)
Disulfide trapping studies have revealed that the pore-lining (P) segments of voltage-dependent sodium channels undergo sizable motions on a subsecond time scale. Such motions of the pore may be necessary for selective ion translocation. Although traditionally viewed as separable properties, gating and permeation are now known to interact extensively in various classes of channels. We have investigated the interaction of pore motions and voltage-dependent gating in micro1 sodium channels engineered to contain two cysteines within the P segments. Rates of catalyzed internal disulfide formation (kSS) were measured in K1237C+W1531C mutant channels expressed in oocytes. During repetitive voltage-clamp depolarizations, increasing the pulse duration had biphasic effects on the kSS, which first increased to a maximum at 200 msec and then decreased with longer depolarizations. This result suggested that occupancy of an intermediate inactivation state (IM) facilitates pore motions. Consistent with the known antagonism between alkali metals and a component of slow inactivation, kSS varied inversely with external [Na+]o. We examined the converse relationship, namely the effect of pore flexibility on gating, by measuring recovery from inactivation in Y401C+E758C (YC/EC) channels. Under oxidative conditions, recovery from inactivation was slower than in a reduced environment in which the spontaneous YC/EC cross-link is disrupted. The most prominent effects were slowing of a component with intermediate recovery kinetics, with diminution of its relative amplitude. We conclude that occupancy of an intermediate inactivation state facilitates motions of the P segments; conversely, flexibility of the P segments alters an intermediate component of inactivation. (+info)
Cerebellar Purkinje cell simple spike discharge encodes movement velocity in primates during visuomotor arm tracking.
(2/2595)
Pathophysiological, lesion, and electrophysiological studies suggest that the cerebellar cortex is important for controlling the direction and speed of movement. The relationship of cerebellar Purkinje cell discharge to the control of arm movement parameters, however, remains unclear. The goal of this study was to examine how movement direction and speed and their interaction-velocity-modulate Purkinje cell simple spike discharge in an arm movement task in which direction and speed were independently controlled. The simple spike discharge of 154 Purkinje cells was recorded in two monkeys during the performance of two visuomotor tasks that required the animals to track targets that moved in one of eight directions and at one of four speeds. Single-parameter regression analyses revealed that a large proportion of cells had discharge modulation related to movement direction and speed. Most cells with significant directional tuning, however, were modulated at one speed, and most cells with speed-related discharge were modulated along one direction; this suggested that the patterns of simple spike discharge were not adequately described by single-parameter models. Therefore, a regression surface was fitted to the data, which showed that the discharge could be tuned to specific direction-speed combinations (preferred velocities). The overall variability in simple spike discharge was well described by the surface model, and the velocities corresponding to maximal and minimal discharge rates were distributed uniformly throughout the workspace. Simple spike discharge therefore appears to integrate information about both the direction and speed of arm movements, thereby encoding movement velocity. (+info)
Neural encoding in orbitofrontal cortex and basolateral amygdala during olfactory discrimination learning.
(3/2595)
Orbitofrontal cortex (OFC) is part of a network of structures involved in adaptive behavior and decision making. Interconnections between OFC and basolateral amygdala (ABL) may be critical for encoding the motivational significance of stimuli used to guide behavior. Indeed, much research indicates that neurons in OFC and ABL fire selectively to cues based on their associative significance. In the current study recordings were made in each region within a behavioral paradigm that allowed comparison of the development of associative encoding over the course of learning. In each recording session, rats were presented with novel odors that were informative about the outcome of making a response and had to learn to withhold a response after sampling an odor that signaled a negative outcome. In some cases, reversal training was performed in the same session as the initial learning. Ninety-six of the 328 neurons recorded in OFC and 60 of the 229 neurons recorded in ABL exhibited selective activity during evaluation of the odor cues after learning had occurred. A substantial proportion of those neurons in ABL developed selective activity very early in training, and many reversed selectivity rapidly after reversal. In contrast, those neurons in OFC rarely exhibited selective activity during odor evaluation before the rats reached the criterion for learning, and far fewer reversed selectivity after reversal. The findings support a model in which ABL encodes the motivational significance of cues and OFC uses this information in the selection and execution of an appropriate behavioral strategy. (+info)
Correlated firing in rabbit retinal ganglion cells.
(4/2595)
A ganglion cell's receptive field is defined as that region on the retinal surface in which a light stimulus will produce a response. While neighboring ganglion cells may respond to the same stimulus in a region where their receptive fields overlap, it generally has been assumed that each cell makes an independent decision about whether to fire. Recent recordings from cat and salamander retina using multiple electrodes have challenged this view of independent firing by showing that neighboring ganglion cells have an increased tendency to fire together within +/-5 ms. However, there is still uncertainty about which types of ganglion cells fire together, the mechanisms that produce coordinated spikes, and the overall function of coordinated firing. To address these issues, the responses of up to 80 rabbit retinal ganglion cells were recorded simultaneously using a multielectrode array. Of the 11 classes of rabbit ganglion cells previously identified, coordinated firing was observed in five. Plots of the spike train cross-correlation function suggested that coordinated firing occurred through two mechanisms. In the first mechanism, a spike in an interneuron diverged to produce simultaneous spikes in two ganglion cells. This mechanism predominated in four of the five classes including the ON brisk transient cells. In the second mechanism, ganglion cells appeared to activate each other reciprocally. This was the predominant pattern of correlated firing in OFF brisk transient cells. By comparing the receptive field profiles of ON and OFF brisk transient cells, a peripheral extension of the OFF brisk transient cell receptive field was identified that might be produced by lateral spike spread. Thus an individual OFF brisk transient cell can respond both to a light stimulus directed at the center of its receptive field and to stimuli that activate neighboring OFF brisk transient cells through their receptive field centers. (+info)
Thapsigargin inhibits a potassium conductance and stimulates calcium influx in the intact rat lens.
(5/2595)
1. An increase in lens cell calcium has long been associated with cortical cataract. Recently, it has been shown that thapsigargin induces a rise in lens cell calcium by release from endoplasmic reticulum stores. The effects of this rise on the optical and membrane characteristics of the lens were studied in the isolated rat lens. 2. The electrical characteristics of the isolated, perifused rat lens were measured using a two-internal microelectrode technique that permits measurement of plasma membrane conductance (Gm), membrane potential (Vm) and junctional conductance in the intact lens. 3. Thapsigargin (1 microM) induced a rapid overall depolarization of Vm that was accompanied by first a decrease and then an increase in Gm. 4. Replacing external Na+ with tetraethylammonium (TEA) abolished the decrease in Gm. However, a transient increase phase was still observed. 5. The changes in conductance were further characterized by measuring 22Na+ and 45Ca2+ influxes into the isolated lens. Thapsigargin (1 microM) induced a transient increase in 45Ca2+, but did not affect Na+ influx. 6. The Ca2+ channel blocker La3+ (10 microM) totally inhibited the thapsigargin-induced Ca2+ influx. It also blocked the increase in Gm observed in control and in Na+-free-TEA medium. In the absence of external calcium, thapsigargin induced a small depolarization in Vm. 7. These data indicate that thapsigargin induces both a decrease in K+ conductance and an increase in Ca2+ conductance. These probably result from release of stored Ca2+ and subsequent activation of store-operated Ca2+ channels (capacitative Ca2+ entry). 8. Thapsigargin application over the time course of these experiments (24 h) had no effect on junctional conductance or on the transparency of the lens. (+info)
Nitric oxide release in penile corpora cavernosa in a rat model of erection.
(6/2595)
1. Nitric oxide (NO) levels were measured in the corpus cavernosum of urethane-anaesthetized rats by using differential normal pulse voltammetry with carbon fibre microelectrodes coated with a polymeric porphyrin and a cation exchanger (Nafion). A NO oxidation peak could be recorded at 650 mV vs. a Ag-AgCl reference electrode every 100 s. 2. This NO signal was greatly decreased by the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME), given by local and systemic routes, and enhanced by the NO precursor L-arginine. Treatment with L-arginine reversed the effect of L-NAME on the NO peak. 3. Both the NO signal and the intracavernosal pressure (ICP) were increased by electrical stimulation of cavernosal nerves (ESCN). However, the rise in the NO levels long outlived the rapid return to baseline of the ICP values at the end of nerve stimulation. 4. The ICP and the NO responses to ESCN were suppressed by local and systemic injections of L-NAME. Subsequent treatment with L-arginine of L-NAME-treated animals restored the NO signal to basal levels and the NO response to ESCN. The ICP response to ESCN was restored only in part by L-arginine. 5. The observed temporal dissociation between the NO and ICP responses could be accounted for by several factors, including the buffering of NO by the blood filling the cavernosal spaces during erection. 6. These findings indicate that an increased production of NO in the corpora cavernosa is necessary but not sufficient for maintaining penile erection and suggest a complex modulation of the NO-cGMP-cavernosal smooth muscle relaxation cascade. (+info)
In vitro simultaneous measurements of relaxation and nitric oxide concentration in rat superior mesenteric artery.
(7/2595)
1. The relationship between nitric oxide (NO) concentration measured with an NO-specific microelectrode and endothelium-dependent relaxation was investigated in isolated rat superior mesenteric artery contracted with 1 microM noradrenaline. 2. Acetylcholine (10 microM) induced endothelium-dependent simultaneous increases in luminal NO concentration of 21 +/- 6 nM, and relaxations with pD2 values and maximum of 6.95 +/- 0.32 and 97.5 +/- 0.7 % (n = 7), respectively. An inhibitor of NO synthase, N G-nitro-L-arginine (L-NOARG, 100 microM) inhibited the relaxations and increases in NO concentration induced by acetylcholine. 3. Oxyhaemoglobin (10 microM) reversed the relaxations and increases in NO concentrations induced by acetylcholine, S-nitroso-N-acetylpenicillamine (SNAP) and S-morpholino-sydnonimine (SIN-1), but not the relaxations induced with forskolin. Oxyhaemoglobin also decreased the NO concentration below baseline level. 4. In the presence of L-NOARG (100 microM), a small relaxation to acetylcholine (10 microM) of noradrenaline-contracted segments was still seen; oxyhaemogobin inhibited this relaxation and decreased the NO concentration by 14 +/- 4 nM (n = 4). 5. The NO concentration-relaxation relationship for acetylcholine resembled that for SNAP and SIN-1 more than for authentic NO. Thus while 7-17 nM NO induced half-maximal relaxations in response to SNAP or SIN-1, 378 +/- 129 nM NO (n = 4) was needed for half-maximal relaxation to authentic NO. 6. The present study provides direct evidence that the relaxation of the rat superior mesenteric artery with the endothelium-dependent vasodilator acetylcholine is correlated to the endogeneous release of NO. The study also suggests that NO mediates the L-NOARG-resistant relaxations in this artery, and that there is a basal NO release. (+info)
Firing properties of single vasoconstrictor neurones in human subjects with high levels of muscle sympathetic activity.
(8/2595)
1. Single-unit recordings were made from 19 postganglionic muscle vasoconstrictor axons via tungsten microelectrodes in the peroneal nerve in seven healthy subjects with many multi-unit sympathetic discharges at rest ('high group', 75 +/- 5 multi-unit bursts per 100 heart beats, mean +/- s.e.m.). The results were compared with previous data from 14 units in subjects with 21 +/- 2 multi-unit bursts per 100 heart beats ('low group'). 2. In the 'high group' the units fired spontaneously in 35 +/- 4 % of all cardiac intervals. One unit only ever fired once per cardiac interval, 14 units (74 %) generated maximally two to three spikes, and four units (21 %) up to four to five spikes. Of those cardiac intervals in which a unit fired, a single spike occurred in 78 %, two spikes in 18 %, three spikes in 4 % and four spikes in less than 1 % of cardiac intervals. Measured as the inverse of all interspike intervals, the mean rate was 0.33 +/- 0.04 Hz and the mean intraburst frequency 22.2 +/- 1.6 Hz. Most results were similar to those in the 'low group', but in the 'low group' heart rate was higher (64.5 vs. 50.4 beats min-1) and mean firing frequency was higher (0.49 +/- 0.06 Hz). 3. During increases of multi-unit burst activity evoked by sustained inspiratory-capacity apnoea the firing probability of nine units in the 'high group' increased from 33 +/- 6 to 56 +/- 3 % of the cardiac intervals. Simultaneously, the incidence of single spikes decreased and the incidence of multiple spikes per cardiac interval increased, resulting in an increase of mean firing frequency from 0. 23 +/- 0.04 Hz at rest to 1.04 +/- 0.14 Hz during the apnoea. 4. We conclude that single muscle vasoconstrictor neurones usually fire only a solitary spike during sympathetic bursts both in subjects with a high and in subjects with a low number of bursts at rest. Presumably, differences in the numbers of bursts are due mainly to differences in firing probability and recruitment of sympathetic fibres. During acute increases of multi-unit activity, both increases in discharge frequency and recruitment of additional neurones contribute to the increased intensity of an individual sympathetic burst. (+info)