Primary afferent-driven presynaptic inhibition of C-fiber inputs to spinal lamina I neurons.

Presynaptic inhibition of primary afferent terminals is a powerful mechanism for controlling sensory information flow into the spinal cord. Lamina I is the major spinal nociceptive projecting area and monosynaptic input from C-fibers to this region represents a direct pathway for transmitting pain signals to supraspinal centers. Here we used an isolated spinal cord preparation to show that this pathway is under control of the afferent-driven GABAergic presynaptic inhibition. Presynaptic inhibition of C-fiber input to lamina I projection and local-circuit neurons is mediated by recruitment of Aß-, Aδ- and C-afferents. C-fiber-driven inhibition of C-fibers functions as a feedforward mechanism, by which the homotypic afferents control sensory information flow into the spinal cord and regulate degree of the primary nociceptive afferent activation needed to excite the second order neurons. The presynaptic inhibition of C-fiber input to lamina I neurons may be mediated by both synaptic and non-synaptic mechanisms, and its occurrence and extent are quite heterogeneous. This heterogeneity is likely to be reflective of involvement of lamina I neurons in diverse circuitries processing specific modalities of sensory information in the superficial dorsal horn. Thus, our results implicate both low- and high-threshold afferents in the modulation of C-fiber input into the spinal cord.

Distinct mechanisms of signal processing by lamina I spino-parabrachial neurons.

Lamina I spino-parabrachial neurons (SPNs) receive peripheral nociceptive input, process it and transmit to the supraspinal centres. Although responses of SPNs to cutaneous receptive field stimulations have been intensively studied, the mechanisms of signal processing in these neurons are poorly understood. Therefore, we used an ex-vivo spinal cord preparation to examine synaptic and cellular mechanisms determining specific input-output characteristics of the neurons. The vast majority of the SPNs received a few direct nociceptive C-fiber inputs and generated one spike in response to saturating afferent stimulation, thus functioning as simple transducers of painful stimulus. However, 69% of afferent stimulation-induced action potentials in the entire SPN population originated from a small fraction (19%) of high-output neurons. These neurons received a larger number of direct Aδ- and C-fiber inputs, generated intrinsic bursts and efficiently integrated a local network activity via NMDA-receptor-dependent mechanisms. The high-output SPNs amplified and integrated the nociceptive input gradually encoding its intensity into the number of generated spikes. Thus, different mechanisms of signal processing allow lamina I SPNs to play distinct roles in nociception.

Neurons in the lateral part of the lumbar spinal cord show distinct novel axon trajectories and are excited by short propriospinal ascending inputs.

The role of spinal dorsal horn propriospinal connections in nociceptive processing is not yet established. Recently described, rostrocaudally oriented axon collaterals of lamina I projection and local-circuit neurons (PNs and LCNs) running in the dorsolateral funiculus (DLF) may serve as the anatomical substrate for intersegmental processing. Putative targets of these axons include lateral dendrites of superficial dorsal horn neurons, including PNs, and also neurons in the lateral spinal nucleus (LSN) that are thought to be important integrator units receiving, among others, visceral sensory information. Here we used an intact spinal cord preparation to study intersegmental connections within the lateral part of the superficial dorsal horn. We detected brief monosynaptic and prolonged polysynaptic excitation of lamina I and LSN neurons when stimulating individual dorsal horn neurons located caudally, even in neighboring spinal cord segments. These connections, however, were infrequent. We also revealed that some projection neurons outside the dorsal grey matter and in the LSN have distinct, previously undescribed course of their projection axon. Our findings indicate that axon collaterals of lamina I PNs and LCNs in the DLF rarely form functional connections with other lamina I and LSN neurons and that the majority of their targets are on other elements of the dorsal horn. The unique axon trajectories of neurons in the dorsolateral aspect of the spinal cord, including the LSN do not fit our present understanding of midline axon guidance and suggest that their function and development differ from the neurons inside lamina I. These findings emphasize the importance of understanding the connectivity matrix of the superficial dorsal horn in order to decipher spinal sensory information processing.

Basic electrical properties of in situ endothelial cells of small pulmonary arteries during postnatal development.

Small pulmonary arteries are the major determinants of pulmonary artery pressure and vascular resistance. Their endothelium modulates pulmonary resistance, remodeling, and blood fluidity. We developed a method that provides access to the luminal surface of small pulmonary arteries of rat and allows the patch-clamp study of electrical properties of in situ endothelium. At birth, the membrane was predominantly permeable for K(+), showing a resting potential of -70 mV. This conductance was not voltage-dependent and was insensitive to standard blockers of K(+) channels such as tetraethylammonium, charybdotoxin, and 4-aminopyridine. The first 22 d of development were accompanied by an additional expression of a Cl(-) conductance, increasing membrane potential to -45 mV. Acidosis reduced K(+) conductance and depolarized the membrane, whereas alkalosis resulted in hyperpolarization. Two-electrode recordings revealed tight electrical coupling (83%) between neighboring cells in the circumferential direction of the artery. The electrotonic length constant for endothelium was 13.3 microm, indicating that most cells in one cross section of a small artery are well coupled. Thus, the resting membrane conductances in small pulmonary artery endothelial cells change with postnatal development and are modulated by pH.

Suppression of potassium conductance by droperidol has influence on excitability of spinal sensory neurons.

BACKGROUND: During spinal and epidural anesthesia with opioids, droperidol is added to prevent nausea and vomiting. The mechanisms of its action on spinal sensory neurons are not well understood. It was previously shown that droperidol selectively blocks a fast component of the Na+ current. The authors studied the action of droperidol on voltage-gated K+ channels and its effect on membrane excitability in spinal dorsal horn neurons of the rat. METHODS: Using a combination of the patch-clamp technique and the "entire soma isolation" method, the action of droperidol on fast-inactivating A-type and delayed-rectifier K+ channels was investigated. Current-clamp recordings from intact sensory neurons in spinal cord slices were performed to study the functional meaning of K+ channel block for neuronal excitability. RESULTS: Droperidol blocked delayed-rectifier K+ currents in isolated somata of dorsal horn neurons with a half-maximum inhibiting concentration of 20.6 microm. The A-type K+ current was insensitive to up to 100 microm droperidol. At droperidol concentrations insufficient for suppression of an action potential, the block of delayed-rectifier K+ channels led to an increase in action potential duration and, as a consequence, to lowering of the discharge frequency in the neuron. CONCLUSIONS: Droperidol blocks delayed-rectifier K+ channels in a concentration range close to that for suppression of Na+ channels. The block of delayed-rectifier K+ channels by droperidol enhances the suppression of activity in spinal sensory neurons at drug concentrations insufficient for complete conduction block.

Differential block of fast and slow inactivating tetrodotoxin-sensitive sodium channels by droperidol in spinal dorsal horn neurons.

BACKGROUND: Dorsal horn neurons of the spinal cord participate in neuronal pain transmission. During spinal and epidural anesthesia, dorsal horn neurons are exposed to local anesthetics and opioids. Droperidol is usually given with opioids to avoid nausea and vomiting. A recently developed method of "entire soma isolation" has made it possible to study directly the action of droperidol on different components of Na+ current in dorsal horn neurons. METHODS: Using a combination of the whole-cell patch-clamp recording from spinal cord slices and the entire soma isolation method, we studied the direct action of droperidol on two types of Na+ currents in dorsal horn neurons of young rats. RESULTS: The tetrodotoxin-sensitive Na+ current in isolated somata consisted of a fast inactivating (tauF, 0.5-2 ms; 80-90% of the total amplitude) and a slow inactivating (tauS, 6-20 ms; 10-20% of the total amplitude) component. Droperidol, at concentrations relevant for spinal and epidural anesthesia, selectively and reversibly suppressed the fast component with a half-maximum inhibiting concentration (IC50) of 8.3 microm. The slow inactivating component was much less sensitive to droperidol; the estimated IC50 value was 809 microm. CONCLUSIONS: Droperidol selectively blocks fast Na+ channels, the fast and slow components of the Na+ current in dorsal horn neurons are carried through pharmacologically distinct types of Na+ channels, and the effects of droperidol differ from those of local anesthetics and tetrodotoxin, which equipotently suppress both components. Droperidol may be suggested as a pharmacologic tool for separation of different types of inactivating tetrodotoxin-sensitive Na+ channel.

Excitability of the soma in central nervous system neurons.

The ability of the soma of a spinal dorsal horn neuron, a spinal ventral horn neuron (presumably a motoneuron), and a hippocampal pyramidal neuron to generate action potentials was studied using patch-clamp recordings from rat spinal cord slices, the "entire soma isolation" method, and computer simulations. By comparing original recordings from an isolated soma of a dorsal horn neuron with simulated responses, it was shown that computer models can be adequate for the study of somatic excitability. The modeled somata of both spinal neurons were unable to generate action potentials, showing only passive and local responses to current injections. A four- to eightfold increase in the original density of Na(+) channels was necessary to make the modeled somata of both spinal neurons excitable. In contrast to spinal neurons, the modeled soma of the hippocampal pyramidal neuron generated spikes with an overshoot of +9 mV. It is concluded that the somata of spinal neurons cannot generate action potentials and seem to resist their propagation from the axon to dendrites. In contrast, the soma of the hippocampal pyramidal neuron is able to generate spikes. It cannot initiate action potentials in the intact neurons, but it can support their back-propagation from the axon initial segment to dendrites.

Effect of bupivacaine on ATP-dependent potassium channels in rat cardiomyocytes.

Bupivacaine induces fatal arrhythmia when accidentally injected i.v. or overdosed, whereas lidocaine is used as an anti-arrhythmic agent. We have suggested recently that the anti-arrhythmic effect of lidocaine may be explained by suppression of ATP-sensitive potassium (KATP) channels. Therefore, it could be argued that different sensitivities of KATP channels to both drugs could be a reason for their different arrhythmic and anti-arrhythmic properties. In this study, we have investigated the direct action of bupivacaine on KATP channels in cardiomyocytes. The effects of bupivacaine on the cardiac KATP channel were investigated using the patch-clamp technique on enzymatically dissociated cardiomyocytes of adult rats. Bupivacaine was applied to the outer side of excised membrane patches using a multiple-barrel perfusion system. Concentration-response curves indicated that bupivacaine blocked the mean current of the KATP channels at a half-maximum inhibiting concentration (IC50) of 29 mumol litre-1, similar to that reported for lidocaine (43 mumol litre-1). Binding of bupivacaine influenced the gating of this channel, but did not reduce the conductance of the open channel. Bupivacaine and lidocaine were equipotent in blocking KATP channels. However, because of its excessive block of the sodium channel in the inactivated state, block of KATP channels by bupivacaine will only enhance its cardiotoxicity.

Spatial distribution of NA+ and K+ channels in spinal dorsal horn neurones: role of the soma, axon and dendrites in spike generation.

Spinal dorsal horn neurones play an important role in processing sensory information received from primary afferent fibers. The application of the patch-clamp technique to thin slices of rat spinal cord has enabled the study of ionic channels in visually identified dorsal horn neurones. The small soma of these neurones isolated from the slice by means of a novel method of 'entire soma isolation' has become a convenient model for investigating the properties and distributions of ionic channels. The present review summarizes results of recent experiments studying different types of voltage-gated Na+ and K+ channels expressed in dorsal horn neurones. Uneven distribution of the channels between the soma. axon and dendrites appears to play a major role in determining the neuronal excitability. The contribution of the soma, axon and dendrites to generation and propagation of the action potentials in central neurones is discussed.

Axonal expression of sodium channels in rat spinal neurones during postnatal development.

1. Postnatal expression of Na+ channels and development of somatic excitability were studied in dorsal horn neurones of rat using patch-clamp recordings from spinal cord slices in combination with the 'entire soma isolation' method. 2. The amplitude of Na+ current in the intact neurone in the slice increased with postnatal development (days 0-39) with a mean rate of 83 pA day-1. 3. The Na+ current in the neuronal soma did not increase with age and the soma, separated from the axon, was not able to fire spikes at any stage of development studied. 4. It is concluded that the postnatal development of the spinal dorsal horn neurone is accompanied by intensive expression of Na+ channels in the axonal but not somatic membrane. The estimated minimum density of Na+ channels in the axon initial segment is approximately 160 channels microm-2.

Na+-activated K+ channels in small dorsal root ganglion neurones of rat.

1. Whole-cell Na+-activated K+ (KNa) channel currents and single KNa channels were studied with the patch-clamp method in small (20-25 micrometer) dorsal root ganglion (DRG) neurones in slices of rat dorsal root ganglia. 2. The whole-cell KNa channel current was identified as an additional K+-selective leakage current which appeared after cell perfusion with internal solutions containing different [Na+]. The concentration for half-maximal activation of KNa channel current was 39 mM and the Hill coefficient was 3.5. At [Na+]i above 12 mM, KNa channel current dominated the unspecific leakage current. The ratio of maximum KNa channel current to unspecific leakage current was 45. 3. KNa channel current was not activated by internal Li+. It was suppressed by external 20 mM Cs+ but not by 10 mM tetraethylammonium. 4. Single KNa channels with a conductance of 142 pS in 155 mM external K+ (K+o)-85 mM internal K+ (K+i) solutions were observed at a high density of about 2 channels micrometer-2. 5. In two-electrode experiments, a direct correlation was seen between development of whole- cell KNa channel current and activation of single KNa channels during perfusion of the neurone with Na+-containing internal solution. 6. Under current-clamp conditions, KNa channels did not contribute to the action potential. However, internal perfusion of the neurone with Na+ shifted the resting potential towards the equilibrium potential for K+ (EK). Varying external [K+] indicated that in neurones perfused with Na+-containing internal solution the resting potential followed the EK values predicted by the Nernst equation over a broader voltage range than in neurones perfused with Na+-free solution. 7. It is concluded that the function of KNa channels has no links to firing behaviour but that the channels could be involved in setting or stabilizing the resting potential in small DRG neurones.

Uneven distribution of K+ channels in soma, axon and dendrites of rat spinal neurones: functional role of the soma in generation of action potentials.

1. A novel method of 'entire soma isolation' was used to describe the distribution of voltage-gated K+ channels between soma, axon and dendrites of dorsal horn neurones identified in spinal cord slices of newborn rat. 2. The soma contained 36 % of total inactivating (KA) current but only 15 % of delayed rectifier (KDR) current. The axon initial segment possessed almost half (47 %) of the total KA current and 38 % of KDR current. In contrast, dendrites contained a small portion (17 %) of KA but 47 % of KDR current. 3. Under current-clamp conditions, the soma isolated from axon and dendrites was not able to generate action potentials. It passively conducted weak (/= 0 mV) depolarizations. 4. It is concluded that the soma plays a complex role in the excitability of spinal dorsal horn neurones. It conducts passively or amplifies excitatory postsynaptic potentials on their way from dendrites and soma to the axon initial segment but it inhibits back-propagation of the action potential from the axon to the dendrites.

Large conductance Ca(2+)-activated K+ channels in the soma of rat motoneurones.

Properties of large conductance Ca(2+)-activated K+ channels were studied in the soma of motoneurones visually identified in thin slices of neonatal rat spinal cord. The channels had a conductance of 82 +/- 5 pS in external Ringer solution (5.6 mM Ko+/(/)155 mM Ki+) and 231 +/- 4 pS in external high-Ko solution (155 mM Ko+/(/)155 mM Ki+). The channels were activated by depolarization and by an increase in internal Ca2+ concentration. Potentials of half-maximum channel activation (E50) were -13, -34, -64 and -85 mV in the presence of 10(-6), 10(-5), 10(-4) and 10(-3) M internal Ca2+, respectively. Using an internal solution containing 10(-4) M Ca2+, averaged KCa currents showed fast activation within 2-3 msec after a voltage step to +50 mV. Averaged KCa currents did not inactivate during 400 msec voltage pulses. External TEA reduced the apparent single-channel amplitude with a 50% blocking concentration (IC50) of 0.17 +/- 0.02 mM. KCa channels were completely suppressed by externally applied 100 mM charybdotoxin. It is concluded that KCa channels activated by Ca2+ entry during the action potential play an important role in the excitability of motoneurones.

Blockade of Na+ and K+ currents by local anesthetics in the dorsal horn neurons of the spinal cord.

BACKGROUND: The dorsal horn of the spinal cord is a pivotal point for transmission of neuronal pain. During spinal and epidural anesthesia, the neurons of the dorsal horn are exposed to local anesthetics. Unfortunately, little is known about the action of local anesthetics on the major ionic conductances in dorsal horn neurons. In this article, the authors describe the effects of bupivacaine, lidocaine, and mepivacaine on voltage-gated Na+ and K+ currents in the membranes of these neurons. METHODS: The patch-clamp technique was applied to intact dorsal horn neurons from laminae I-III identified in 200-microm slices of spinal cord from newborn rats. Under voltage-clamp conditions, the whole-cell Na+ and K+ currents activated by depolarization were recorded in the presence of different concentrations of local anesthetics. RESULTS: Externally applied bupivacaine, lidocaine, and mepivacaine produced tonic block of Na+ currents with different potencies. Half-maximum inhibiting concentrations (IC50) were 26, 112, and 324 microM, respectively. All local anesthetics investigated also showed a phasic, that is, a use-dependent, block of Na+ channels. Rapidly inactivating K+ currents (KA currents) also were sensitive to the blockers with IC50 values for tonic blocks of 109, 163, and 236 microM, respectively. The block of KA currents was not use dependent. In contrast to Na+ and KA currents, delayed-rectifier K+ currents were almost insensitive to the local anesthetics applied. CONCLUSIONS: In clinically relevant concentrations, local anesthetics block Na+ and KA currents but not delayed-rectifier K+ currents in spinal dorsal horn neurons. The molecular mechanisms of Na+ and K+ channel block by local anesthetics seem to be different. Characterization of these mechanisms could be an important step in understanding the complexity of local anesthetic action during spinal and epidural anesthesia.

Functional distribution of three types of Na+ channel on soma and processes of dorsal horn neurones of rat spinal cord.

1. Voltage-gated Na+ channels and their distribution were studied by the patch-clamp technique in intact dorsal horn neurones in slices of newborn rat spinal cord and in neurones isolated from the slice by slow withdrawal of the recording pipette. This new method of neurone isolation was further used to study the roles of soma and axon in generation of action potentials. 2. Tetrodotoxin (TTX)-sensitive Na+ currents in intact neurones consisted of three components. A fast component with an inactivation time constant (tau f) of 0.6-2.0 ms formed the major part (80-90%) of the total Na+ current. The remaining parts consisted of a slowly inactivating component (tau s of 5-20 ms) and a steady-state component. 3. Single fast and slow inactivating Na+ channels with conductances of 11.6 and 15.5 pS, respectively, were identified in the soma of intact neurones in the slice. Steady-state Na+ channels were not found in the soma, suggesting an axonal or dendritic localization of these channels. 4. In the whole-cell recording mode, the entire soma of a dorsal horn neurone could be isolated from the slice by slow withdrawal of the recording pipette, leaving all or nearly all of its processes in the slice. The isolated structure was classified as: (1) 'soma' if it lost all of its processes, (2) 'soma+axon' complex if it preserved one process and at least 85% of its original peak Na+ current or (3) 'soma+dendrite' complex if it preserved one process but the remaining Na+ current did not exceed those observed in the isolated 'somata'. 5. The spatial distribution of Na+ channels in the neurone was studied by comparing Na+ currents recorded before and after isolation. The isolated 'soma' contained 13.8 +/- 1.3% of inactivating Na+ current but no steady-state Na+ current. 'Soma+axon' complexes contained 93.6 +/- 1.4% of inactivating and 46% of steady-state Na+ current. 6. In current-clamp experiments, the intact neurones and isolated 'soma+axon' complexes responded with 'all-or-nothing' action potentials to current injections. In contrast, isolated 'somata' showed only passive or local responses and were unable to generate action potentials. 7. It is concluded that dorsal horn neurones of the spinal cord possess three types of TTX-sensitive voltage-gated Na+ channels. The method of entire soma isolation described here shows that the majority of inactivating Na+ channels are localized in the axon hillock and only a small proportion (ca 1/7) are distributed in the soma. Steady-state Na+ channels are most probably expressed in the axonal and dendritic membranes. The soma itself is not able to generate action potentials. The axon or its initial segment plays a crucial role in the generation of action potentials.

Properties and functions of Na(+)-activated K+ channels in the soma of rat motoneurones.

1. Properties and functions of Na(+)-activated K+ (KNa) channels in the soma of motoneurones were studied in spinal cord slices of newborn rat. KNa channels had a conductance of 44.8 pS in 5.6 mM external K+ (Ko+)/106 mM internal K+ (Ki+) solutions and 139.2 pS in 155 mM Ko+/85 mM Ki+ solutions. KNa channels were voltage independent and needed a relatively high [Na+]i to become active (EC50 = 39.9 mM). Li+ could not substitute for Na+ in activation of KNa channels. The channels were predominantly found in the vicinity of cell processes, in the regions of most probable accumulation of cytoplasmic Na+. 2. In current-clamp experiments, the shape of the single action potential (AP) recorded in Ca(2+)-free Ringer solution was not changed after substitution of external Na+ with Li+. However, 0.4-0.8 s trains of APs were followed by a slow (1-2s) after-hyperpolarization (sAHP), which reversibly disappeared when external Na+ was replaced by Li+. Na(+)-activated sAHP persisted after addition of ouabain and its amplitude was even increased in K(+)-free Ringer solution. sAHP disappeared when the membrane potential was equal to the K+ equilibrium potential. This indicated that sAHP resulted from activation of a Na(+)-dependent K+ conductance, rather than from activation of the electrogenic Na(+)-K+ pump. 3. In conclusion, KNa channels can play an important role in excitability of motoneurones. KNa channels do not make a contribution to the single AP, but they can be activated by a local accumulation of internal Na+ during trains of APs. A Na(+)-activated K+ conductance can reduce membrane excitability and contribute to regulation of AP firing in motoneurones.

Single voltage-gated K+ channels and their functions in small dorsal root ganglion neurones of rat.

1. Single voltage-activated K+ channels were investigated by means of the patch-clamp technique in small dorsal root ganglion (DRG) neurones in 150 microns thin slices of new-born rat DRG. It was found that K+ conductance in small DRG neurones is formed by one type of fast inactivating A-channel and four types of delayed rectifier K+ channels, which could be separated on the basis of their single-channel conductance, kinetics and sensitivity to external tetraethylammonium (TEA). 2. Potassium A-channels were observed at relatively moderate density. They were weakly sensitive to TEA and activated between -70 and +20 mV. The conductance of A-channels was about 40 pS for inward currents in symmetrical high-K+ solutions with external 5 mM TEA added to suppress other types of K+ channels. The time constant of channel inactivation (tau in) was 18.8 ms at -70 mV and 6 ms at potentials positive to -20 mV. 3. A fast delayed rectifier (DRF) channel with a conductance of 55 pS in symmetrical high-K+ solutions was the most frequent type of K+ channel. The channel activated in a broad potential range between -50 and +60 mV and demonstrated a fast deactivation within 1-3 ms after potential return to -80 mV in high-Ko+ solution. The tau in value was 90-150 ms at positive membrane potentials. The single-channel current amplitudes were blocked to 55% by 1 mM TEA. 4. Three further types of delayed rectifier K+ channels were called DR1-, DR2- and DR3- channels. Their single-channel conductances for inward currents in symmetrical high-K+ solutions were distributed between 30 and 44 pS. The channels activated in almost the same voltage range between -60 and -10 mV. Deactivation of the channels at -80 mV lasted tens of milliseconds. The channels were separated on the basis of their sensitivities to TEA. DR1-channel currents were reduced to 50% in the presence of 1 mM TEA, DR2-channel currents were reduced to about 50% by 5 mM TEA, whereas the amplitudes of currents through DR3-channels were almost unaffected by 5 mM TEA. 5. Addition of external 1 and 5 mM TEA to whole cells under current-clamp condition depolarized the cell membrane, lowered the threshold for action potential firing, prolonged action potential duration and reduced the amplitude of after-hyperpolarization. 6. It is concluded that potassium A-, DRF-, DR1-, DR2- and DR3-channels play multiple roles in the excitability of DRG neurones. Possible influences of these channels on the shape of the action potential, its firing threshold and the resting membrane potential of small DRG neurones are discussed.

Modulation of delayed rectifier K+ channel activity by external K+ ions in Xenopus axon.

The effect of external K+ ions upon the activation of delayed rectifier K+ channels was studied in demyelinated amphibian nerve fibres by means of the patch-clamp technique. In external 105 mM K+ solution (high-Ko) macroscopic K+ currents activated at more negative potentials (approximately -15 mV) than in external Ringer (2.5 mM K+). Since the rapid substitution of external Ringer with high-Ko solution at holding potentials of -70 mV and -60 mV directly activated K+ concentration from 5 mM to 10,20,50 and 105 mM gradually increased the open probability of the channels. Although Rb+ ions were less permeant through the channels, they were more potent in their interaction with the binding site and shifted K+ channel activation to more negative potentials. In contrast, external Cs+ ions had only a weak effect on the binding site. Thus, external K+ ions at physiological concentrations modulate the activation of delayed rectifier K+ channels at potentials between -90 mV and -60 mV.

Single voltage-activated Na+ and K+ channels in the somata of rat motoneurones.

1. Voltage-activated Na+ and K+ channels were investigated in the soma membrane of motoneurones using the patch-clamp technique applied to thin slices of neonatal rat spinal cord. 2. One type of TTX-sensitive Na+ channel, with a conductance of 14.0 pS, was found to underlie the macroscopic Na+ conductance in the somata of motoneurones. These channels activated within a potential range between -60 and -20 mV with a potential of half-maximal activation (E50) of -38.9 mV and steepness factor (k) of 6.1 mV. 3. Kinetics of Na+ channel inactivation could be fitted with a single exponential function at all potentials investigated. The curve of the steady-state inactivation had the following parameters: a half-maximal potential (Eh,50) of -81.6 mV and k of -10.2 mV. 4. Kinetics of recovery of Na+ channels from inactivation at a potential of -80 mV were double exponential with fast and slow components of 16.2 (76%) and 153.7 ms (24%), respectively. It is suggested that the recovery of Na+ channels from inactivation plays a major role in defining the limiting firing frequency of action potentials in motoneurones. 5. Whole-cell K+ currents consisted of transient (A)- and delayed-rectifier (DR)-components. The A-component activated between -60 and +20 mV with an E50 of -33.3 mV and k of 15.7 mV. The curve of steady-state inactivation was best fitted with an Eh,50 of -82.5 mV and k of -10.2 mV. The DR-component of K+ current activated smoothly at more positive potentials. E50 and k for DR-currents were +1.4 and 16.9 mV, respectively. 6. The most frequent single K+ channel found in the somata of motoneurones was the fast inactivating A-channel with a conductance of 19.2 pS in external Ringer solution. In symmetrical high-K+ solutions the conductance was 50.9 and 39.6 pS for inward and outward currents, respectively. The channel activation took place between -60 and +20 mV. The curve of steady-state inactivation of single A-channels had an Eh,50 of -87.1 mV and k of -12.8 mV. In high-Ko+ solution A-channels demonstrated a rapid deactivation at potentials between -110 and -60 mV. The time constant of the channel deactivation depended on the membrane potential and changed from 1.5 ms at -110 mV to 6.3 ms at -60 mV. 7. Delayed-rectifier K+ channels were found in the soma membrane at a moderate density.(ABSTRACT TRUNCATED AT 250 WORDS)