Block of sodium currents in rat dorsal root ganglion neurons by diphenhydramine

To elucidate the local anesthetic mechanism of diphenhydramine, its effects on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium currents in rat dorsal root ganglion (DRG) neurons were examined by the whole-cell voltage clamp method. Diphenhydramine blocked TTX-S and TTX-R sodium currents with K(d) values of 48 and 86 microM, respectively, at a holding potential of -80 mV. It shifted the conductance-voltage curve for TTX-S sodium currents in the depolarizing direction but had little effect on that for TTX-R sodium currents. Diphenhydramine caused a shift of the steady-state inactivation curve for both types of sodium currents in the hyperpolarizing direction. The time-dependent inactivation became faster and the recovery from the inactivation was slowed by diphenhydramine in both types of sodium currents. Diphenhydramine produced a profound use-dependent block when the cells were repeatedly stimulated with high-frequency depolarizing pulses. The use-dependent block was more pronounced in TTX-R sodium currents. The results show that diphenhydramine blocks sodium channels of sensory neurons similarly to local anesthetics.     

N-Ethylmaleimide modulation of tetrodotoxin-sensitive and tetrodotoxin-resistant sodium channels in rat dorsal root ganglion neurons

The effects of N-ethylmaleimide (NEM), an alkylating reagent to protein sulfhydryl groups, on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium channels in rat dorsal root ganglion (DRG) neurons were studied using the whole cell configuration of patch-clamp technique. When currents were evoked by step depolarizations to 0 mV from a holding potential of −80 mV NEM decreased the amplitude of TTX-S sodium current, but exerted little or no effect on that of TTX-R sodium current. The inhibitory effect of NEM on TTX-S sodium channel was mainly due to the shift of the steady-state inactivation curve in the hyperpolarizing direction. NEM did not affect the voltage-dependence of the activation of TTX-S sodium channel. The steady-state inactivation curve for TTX-R sodium channel was shifted by NEM in the hyperpolarizing direction as that for TTX-S sodium channel. NEM caused a change in the voltage-dependence of the activation of TTX-R sodium channel unlike TTX-S sodium channel. After NEM treatment, the amplitudes of TTX-R sodium currents at test voltages below −10 mV were increased, but those at more positive voltages were not affected. This was explained by the shift in the conductance–voltage curve for TTX-R sodium channels in the hyperpolarizing direction after NEM treatment.     

Modulation of sodium currents in rat dorsal root ganglion neurons by sulfur dioxide derivatives

The effect of sulfur dioxide (SO2) derivatives, a common air pollutant and exists in vivo as an equilibrium between bisulfate and sulfite, on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium channels in cultured post-natal dorsal root ganglion (DRG) neurons were studied using the whole cell configuration of patch-clamp technique. SO2 derivatives on two types of sodium currents were either inhibitory or stimulatory depending on the kinetic parameters tested. At a holding potential of -80 mV, SO2 derivatives suppressed TTX-S sodium currents when depolarizing potential was negative to -30 mV and TTX-R sodium currents when negative to -10 mV but they increased them when the depolarizing potential was positive to -30 or -10 mV. SO2 derivatives shifted the conductance-voltage curve for TTX-R sodium currents in the depolarizing direction but had little effect on that for TTX-S sodium currents. The steady-state inactivation curve for TTX-R sodium channel was shifted by SO2 derivatives in the depolarizing direction as that for TTX-S sodium channel. SO2 derivatives changed the reversal potential and increased the maximum conductance of two types of sodium channels. SO2 derivatives postponed the activating time and delayed the inactivation of sodium currents. The results suggest that SO2 derivatives would increase the excitability of neurons and alter the ion selectivity for two types of sodium currents.     

ATP modulation of sodium currents in rat dorsal root ganglion neurons

The modulation of tetrodotoxin-sensitive (TTX-S) and slow tetrodotoxin-resistant (TTX-R) sodium currents in rat dorsal root ganglion neurons by ATP was studied using the whole-cell patch-clamp method. The effects of ATP on two types of sodium currents were either stimulatory or inhibitory depending on the kinetic parameters tested. At a holding potential of -80 mV ATP suppressed TTX-S sodium currents when the depolarizing potential was positive to -30 mV but it increased them when the depolarizing potential was negative to -30 mV. At the same holding potential slow TTX-R sodium currents were always increased by ATP regardless of the depolarizing potential. In both types of sodium currents ATP shifted both the conductance-voltage relationship curve and the steady-state inactivation curve in the hyperpolarizing direction, and accelerated the time-dependent inactivation. ATP decreased the maximum conductance of TTX-S sodium currents but increased that of slow TTX-R sodium currents. The results suggest that ATP would decrease the excitability of neurons with TTX-S sodium channels but would increase that of neurons with slow TTX-R sodium channels. The effects of ATP on sodium currents were preserved in the presence of a G-protein inhibitor, GDP-beta-S, or purinergic antagonists, suramin and Reactive Blue-2, suggesting that purinergic receptors might not be involved in ATP modulation of sodium currents.     

Modulation of tetrodotoxin-resistant sodium channels by dihydropyrazole insecticide RH-3421 in rat dorsal root ganglion neurons

The effects of the dihydropyrazole insecticide RH-3421 on the retrodotoxin-resistant (TTX-R) voltage-gated sodium channels in rat dorsal root ganglion (DRG) neurons were studied using the whole-cell patch clamp technique. RH-3421 at 10 nM to 1 microM completely blocked action potentials. The sodium currents were irreversibly suppressed by 1 microM RH-3421 in a time- and a dose-dependent manner and the IC50 value of RH-3421 was estimated to be 0.7 microM after 10 min of application. RH-3421 blocked the sodium currents to the same extent over the entire range of test potentials. The sodium conductance-voltage curve was not shifted along the voltage axis by 1 microM RH-3421 application In contrast, both fast and slow steady-state sodium channel inactivation curves were shifted in the hyperpolarizing direction in the presence of 1 microM RH-3421. It was concluded that RH-3421 bound to the resting and inactivated sodium channels to cause block with a higher affinity for the latter state.     

N-Ethylmaleimide modulation of tetrodotoxin-sensitive and tetrodotoxin-resistant sodium channels in rat dorsal root ganglion neurons

The effects of N-ethylmaleimide (NEM), an alkylating reagent to protein sulfhydryl groups, on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium channels in rat dorsal root ganglion (DRG) neurons were studied using the whole cell configuration of patch-clamp technique. When currents were evoked by step depolarizations to 0 mV from a holding potential of -80 mV NEM decreased the amplitude of TTX-S sodium current, but exerted little or no effect on that of TTX-R sodium current. The inhibitory effect of NEM on TTX-S sodium channel was mainly due to the shift of the steady-state inactivation curve in the hyperpolarizing direction. NEM did not affect the voltage-dependence of the activation of TTX-S sodium channel. The steady-state inactivation curve for TTX-R sodium channel was shifted by NEM in the hyperpolarizing direction as that for TTX-S sodium channel. NEM caused a change in the voltage-dependence of the activation of TTX-R sodium channel unlike TTX-S sodium channel. After NEM treatment, the amplitudes of TTX-R sodium currents at test voltages below -10 mV were increased, but those at more positive voltages were not affected. This was explained by the shift in the conductance-voltage curve for TTX-R sodium channels in the hyperpolarizing direction after NEM treatment.     

Sodium currents in striatal neurons from dystonic dt(sz) hamsters: altered response to lamotrigine

Dystonic mutant dt(sz) hamsters are a model for paroxysmal dystonia. Handling/stress provoke the dystonic attacks. This phenomenon subsedes with maturation, but can be reinvoked when these animals receive sodium channel blockers such as lamotrigine, suggesting a dysfunction of striatal sodium channels. Voltage-gated fast sodium currents (I(Na(+))) were studied in acutely isolated striatal neurons from healthy and dt(sz) hamsters in whole-cell voltage clamp recordings. The action of lamotrigine was tested on (a) current/voltage relationship, (b) kinetics, and (c) steady-state inactivation and activation. Under control conditions, properties of I(Na(+)) were not different between healthy and dt(sz) neurons. With lamotrigine, however, (a) peak currents were significantly less depressed by the drug in neurons from dt(sz) hamsters as compared to healthy cells, and (b) the steady-state inactivation curve shift of I(Na(+)) was less pronounced in dt(sz) neurons. The results suggest that in dt(sz) hamsters, fast sodium currents in striatal neurons are more resistant to blockade. This sodium channel alteration might be causal for a functional imbalance between input and output structures of the basal ganglia under conditions of compromised I(+)(Na).     

Resveratrol inhibits Na+ currents in rat dorsal root ganglion neurons

Resveratrol, a phytoalexin found in grapevines, exerts neuroprotective, cancer chemopreventive, antiinflammatory and cardioprotective activities. Studies have also shown that resveratrol exhibits analgesic effects. Cyclooxygenase inhibition and K+ channel opening have been suggested as underlying mechanisms for the resveratrol-induced analgesia. Here, we investigated the effects of resveratrol on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) Na+ currents in rat dorsal root ganglion (DRG) neurons. Resveratrol suppressed both Na+ currents evoked at 0 mV from -80 mV. TTX-S Na+ current (K(d), 72 microM) was more susceptible to resveratrol than TTX-R Na+ current (K(d), 211 microM). Although the activation voltage of TTX-S Na+ current was shifted in the depolarizing direction by resveratrol, that of TTX-R Na+ current was not. Resveratrol caused a hyperpolarizing shift of the steady-state inactivation voltage and slowed the recovery from inactivation of both Na+ currents. However, no frequency-dependent inhibition of resveratrol on either type of Na+ current was observed. The suppression and the unfavorable effects on the kinetics of Na+ currents in terms of the excitability of DRG neurons may make a great contribution to the analgesia by resveratrol.     

Lidocaine stabilizes the open state of CNS voltage-dependent sodium channels

We have previously reported that the lidocaine action is different between CNS and muscle batrachotoxin-modified Na+ channels [Salazar et al., J. Gen. Physiol. 107 (1996) 743-754; Brain Res. 699 (1995) 305-314]. In this study we examined lidocaine action on CNS Na+ currents, to investigate the mechanism of lidocaine action on this channel isoform and to compare it with that proposed for muscle Na+ currents. Na+ currents were measured with the whole cell voltage clamp configuration in stably transfected cells expressing the brain alpha-subunit (type IIA) by itself (alpha-brain) or together with the brain beta(1)-subunit (alphabeta(1)-brain), or the cardiac alpha-subunit (hH1) (alpha-cardiac). Lidocaine (100 microM) produced comparable levels of Na+ current block at positive potentials and of hyperpolarizing shift of the steady-state inactivation curve in alpha-brain and alphabeta(1)-brain Na+ currents. Lidocaine accelerated the rates of activation and inactivation, produced an hyperpolarizing shift in the steady-state activation curve and increased the current magnitude at negative potentials in alpha-brain but not in alphabeta(1)-brain Na+ currents. The lidocaine action in alphabeta(1)-brain resembled that observed in alpha-cardiac Na+ currents. The lidocaine-induced increase in current magnitude at negative potentials and the hyperpolarizing shift in the steady-state activation curve of alpha-brain, are novel effects and suggest that lidocaine treatment does not always lead to current reduction/block when it interacts with Na+ channels. The data are explained by using a modified version of the model proposed by Vedantham and Cannon [J. Gen. Physiol., 113 (1999) 7-16] in which we postulate that the difference in lidocaine action between alpha-brain and alphabeta(1)-brain Na+ currents could be explained by differences in the lidocaine action on the open channel state.     

Effects of lesions of various brain areas on drug priming or footshock-induced reactivation of extinguished conditioned place preference

The effect of Radix paeoniae rubra (RPR) on voltage-gated sodium channel (VGSC) currents (I_Na) was examined in freshly isolated rat hippocampal CA1 neurons using whole-cell patch-clamp technique under voltage-clamp conditions. RPR suppressed I_Na without affecting the current activation, inactivation and deactivation. The amplitude of I_Na decreased by 18.4% within a few seconds of 0.8 mg/ml RPR exposure. RPR (0.8 mg/ml) shifted the steady-state inactivation curves of I_Na to negative potentials, with hyperpolarizing direction shift of V_1/2 of 10.0 mV. The time course of I_Na recovery from inactivation was prolonged significantly by 0.8 mg/ml RPR. RPR (0.8 mg/ml) also enhanced the activity-dependent attenuation of I_Na and decreased the fraction of activated channels. These results suggested that RPR suppressed hippocampal CA1 I_Na by shifting the inactivation curve in hyperpolarizing direction, slowing the recovery time course from inactivation, enhancing the activity-dependent attenuation and decreasing the number of activatable channels. RPR suppression on I_Na might predict the protective effect during brain ischemia in hippocampal CA1 neurons.     

The effects of pumiliotoxin-B on sodium currents in guinea pig hippocampal neurons

The actions of pumiliotoxin-B, extracted from the skin of the frog Dendrobates pumilio, were examined on hippocampal slices and on acutely dissociated hippocampal neurons from the adult guinea pig. Application of 0.5-1 microM pumiliotoxin-B to hippocampal slices caused spontaneous, repetitive field discharges in the CA3 subfield. In whole-cell patch-clamp recordings of isolated CA1 and CA3 neurons, 1-2 microM pumiliotoxin-B shifted the midpoint of Na+ current activation by -11.4 +/- 1.1 mV. This shift was not dependent upon prior activation of the sodium channel. Pumiliotoxin-B did not block macroscopic Na+ inactivation but did reduce the apparent voltage-dependence of inactivation such that currents decayed faster at membrane potentials more negative than -30 mV. Single-channel recordings of sodium currents from excised membrane patches indicated that pumiliotoxin-B had little or no effect on channel closings due to entry into inactivated state(s) but did increase the rate of channel closings due to reversal of channel opening. The increase in the channel closing rate was consistent with a +8.7 mV shift in voltage sensitivity. Negative shifts in activation and positive shifts in closing rates implied a negative shift in the voltage-dependence of channel opening, suggesting that pumiliotoxin-B increases the rate of Na+ channel opening and closing in cells at rest, which could result in spontaneous activity in the neurons.     

Differential effects of K(+) channel blockers on frequency-dependent action potential broadening in supraoptic neurons

Recordings were made from magnocellular neuroendocrine cells dissociated from the supraoptic nucleus of the adult guinea pig to determine the role of voltage gated K(+) channels in controlling the duration of action potentials and in mediating frequency-dependent action potential broadening exhibited by these neurons. The K(+) channel blockers charybdotoxin (ChTx), tetraethylammonium (TEA), and 4-aminopyridine (4-AP) increased the duration of individual action potentials indicating that multiple types of K(+) channel are important in controlling action potential duration. The effect of these K(+) channel blockers was almost completely reversed by simultaneous blockade of voltage gated Ca(2+) channels with Cd(2+). Frequency-dependent action potential broadening was exhibited by these neurons during trains of action potentials elicited by membrane depolarizing current pulses presented at 10 Hz but not at 1 Hz. 4-AP but not ChTx or TEA inhibited frequency-dependent action potential broadening indicating that frequency-dependent action potential broadening is dependent on increasing steady-state inactivation of A-type K(+) channels (which are blocked by 4-AP). A model of differential contributions of voltage gated K(+) channels and voltage gated Ca(2+) channels to frequency-dependent action potential broadening, in which an increase of Ca(2+) current during each successive action potential is permitted as a result of the increasing steady-state inactivation of A-type K(+) channels, is presented.     

Voltage-dependent sodium and potassium currents in cultured trout astrocytes

Voltage-gated ionic currents were recorded from cultured trout astrocytes with the whole-cell variation of the patch-clamp technique. In a subpopulation of astrocytes depolarizations above ?40 m V activated a fast transient inward current that was identified as a sodium current by ion substitution experiments, its current reversal potential, and its TTX-sensitivity. Regarding threshold of activation, peak current voltage, and amplitude this current closely resembled those previously described for mammalian astrocytes. Voltage-dependence of inactivation and kinetics, however, markedly differed from the “glial-like” sodium current occurring in mammalian hippocampal or optic nerve astrocytes, since the sodium current of trout astrocytes exhibited a faster time course of activation and decay and a more depolarized steady-state inactivation curve with midpoints close to ?60 mV. During a period of 2 weeks in culture the biophysical properties of the sodium current did not change significantly, albeit a continuous decrease in current density was observed. At depolarizing voltage steps positive to ?40 mV, additionally voltage-gated potassium outward currents were evoked, which could be separated into a steady-state current with delayed rectifier properties and an inactivating component resembling the A-type current. Moreover, in a subpopulation of astrocytes an inward potassium current was elicited at hyperpolarizing potentials, which exhibited biophysical features consistent with the potassium inward rectifier of mammalian astrocytes. © 1994 Wiley-Liss, Inc.     

Lamotrigine Reduces Voltage-Gated Sodium Currents in Rat Central Neurons in Culture

Summary: Purpose : To study the mechanism or mechanisms of action of lamotrigine (LTG) and, in particular, to establish its effects on the function of NA⁺ channels in mammalian central neurons.
Methods : Rat cerebellar granule cells in culture were subjected to the whole-cell mode of voltage clamping under experimental conditions designed to study voltage-gated Na⁺ currents.
Results : Extracellular application of LTG (10–500 μ M , n = 21) decreased in a dose-related manner a tetrodotoxin-sensitive inward current that was elicited by depolarizing commands (from −80 to +20mV). The peak amplitude of this Na⁺-mediated current was diminished by 38.8 ± 12.2% (mean ± SD, n = 6) during application of 100 μ M LTG, and the dose-response curve of this effect indicated an IC₅₀ 145 μM. The reduction in the inward currents produced by LTG was not associate with any signficant change in the current decay, whereas the voltage dependency of the steady-state inactivation shifted toward more negative values (midpoint of the inactivation curve: –47.5 and –59.0 mV under control conditions and during application of 100 μM LTG, respectively, n = 4).
Conclusions : Our findings indicate that LTG reduces the amplitude of voltage-gated Na⁺ inward current in rat cerebellar granule cells and induces a negative shift of the steady-state inactivation curve. Both mechanisms may be instrumental in controlling the repetitive firing of action potentials (AP) that occurs in neuronal networks during seizure activity.     

Rundown of a transient potassium current is attributable to changes in channel voltage dependence

Many ionic currents undergo significant rundown during whole-cell recording. Although rundown is an artifact associated with the recording method, studying the mechanism of rundown may lead to understanding mechanisms regulating channel functions in physiological conditions. The mechanisms for rundown, however, remain obscure for many channels. Here we have studied the mechanism for rundown of an A-type K(+) current in mouse striatal cholinergic interneurons. The interneuron expressed a prominent component of A-type current which exhibited significant rundown during whole-cell recording. When the current was assessed with a highly hyperpolarized prepotential (-140 mV), however, the rundown was virtually fully suppressed, suggesting its being dependent on voltage. Estimation of channel voltage dependence revealed that both activation and inactivation curves shifted towards hyperpolarized potentials during rundown. The shift was suppressed by intracellular ATP, but was affected neither by phosphatase inhibitors nor by antioxidative reagents. The gradual shift of inactivation curve towards negative potentials would make the holding potential progressively inactivate the channel, resulting in apparent loss of activity of the channels. Our results thus provide a biophysical explanation for rundown of A-type current. .     

Voltage-gated sodium channels expressed in the human cerebellar medulloblastoma cell line TE671

A characterization of the properties of voltage-gated sodium channels expressed in the human cerebellar medulloblastoma cell line TE671 is presented. Membrane currents were recorded under voltage clamp conditions using the patch clamp technique in both the whole-cell and the excised-patch configurations. Macroscopic sodium currents display a typical transient time course with a sigmoidal rise to a peak followed by an exponential decay. The rates of early activation and subsequent inactivation accelerate and approach a maximum in response to test potentials, V, of greater depolarization. The magnitude of peak sodium current increased from negligible values below V = -50 mV and reached a maximum at V = -3.6 mV +/- 2.7 mV (mean +/- S.E.M., n = 12). Sodium currents reversed at V = + 70 mV, near the predicted Nernst equilibrium potential for a Na+ selective channel. The peak sodium conductance, gpeak increased with depolarizing voltages to a maximum at V = approximately 0 mV, exhibiting half-activation voltage at V approximately equal to -36.8 mV and an e-fold change in gpeak/9.5 mV. The Hodgkin-Huxley inactivation parameter h infinity indicates that at V = -73.6 mV half of the sodium currents were inactivated. Single channel current recordings demonstrated the occurrence of discrete events: the latency for first opening was shorter as the depolarizing pulse became more positive. The single-channel current amplitude was ohmic with a slope conductance, gamma = 17.13 pS +/- 0.66 pS. Sodium channel currents were reversibly blocked by tetrodotoxin (TTX).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of pentylenetetrazol on molluscan neurons II. Voltage clamp studies

The effects of pentylenetetrazol (PTZ) upon the steady and transient outward ionic currents during PTZ-induced prolonged depolarizations were investigated using voltage clamp techniques. PTZ causes a 5–35% reduction in g_L and a 40–60% reduction in steady-state g_K. There is also a marked reduction in the activation of g_A of Connor and Stevens⁶ at all clamp potentials; a shortening of the time constant for the inactivation of g_A; and a 10–15 mV shift in the depolarizing direction of the curve relating the steady-state inactivation of g_A to membrane potential. The equillibrium potentials for both g_A and g_K are depolarized by 20 mV in PTZ solution. Equation and voltage clamp data for normal repetitive firing were integrated with the normal and PTZ-altered data. Solution to these equations demonstrated: (1) normal repetitive firing in response to a constant current stimulus; and (2) PTZ-altered repetitive firing that was in the direction of, and for the most part, similar to the observed behavior.     

Oxidative modulation of the transient potassium current IA by intracellular arachidonic acid in rat CA1 pyramidal neurons

Oxidative stress affects cellular membrane lipids and proteins. Using whole-cell patch-clamp recording we demonstrate differential oxidative inhibition of voltage-gated transient (IA) and delayed rectifier [IK(V)] K+ currents by arachidonic acid (AA) and H2O2 in CA1 neurons in hippocampal slice. We show that intracellular application of 1 pm AA or its non-metabolizable analog eicosatetraynoic acid (100 pm) reduced IA by approximately 42% but did not affect IK(V). AA shifted the voltage dependence of steady-state inactivation of IA by 12 mV to more negative potentials whereas the rate of inactivation was unchanged. Surprisingly, intracellular glutathione (GSH, 20 mm) enhanced the effect of AA on maximal IA (-62%) and with AA slowed inactivation of IA. The combination of GSH and extracellular ascorbate (0.4 mm) prevented reduction of IA by AA. Intracellular Trolox (a vitamin E analog, 10 microm) reduced IA by 61%and IK(V) by 39%. Like AA, intracellular Trolox caused a 10-mV left shift of IA steady-state inactivation but Trolox and AA did not cause a shift when coapplied. Extracellular Trolox (100 microm) had no effects on IA. H2O2 (80 microm) reduced both IA and IK(V) in a GSH- and ascorbate-sensitive manner and slowed the rate of inactivation of IA by a factor of 2. Coapplication of H2O2 with GSH and extracellular ascorbate caused approximately 22 mV negative shifts of both steady-state inactivation and activation. We conclude that AA is extremely potent in affecting IA by oxidative modifications. Antioxidants can augment these effects, probably by catalysis of the underlying reactions between oxidants and IA channel proteins.     

Effects of methylmercury on electrical responses of neuroblastoma cells

The effects of methylmercury on a variety of electrophysiological properties of the N1E-115 neuroblastoma cells were studied using microelectrode and voltage clamp techniques. The action potential was reduced in amplitude with an apparent dissociation constant of the order of 20 microM in the face of relatively small membrane depolarization. Voltage clamp experiments revealed that both peak sodium current and steady-state potassium current were suppressed by 20-60 microM methylmercury, with a stronger effect on the sodium current than on the potassium current. The protein reagents dithiodipyridine and N-ethylmaleimide suppressed both currents. Acetylcholine receptor/channel complexes are vulnerable to the action of methylmercury; the nicotinic fast depolarizing response, the muscarinic hyperpolarizing response, and the muscarinic slow depolarizing response, were all suppressed by 10-30 microM methylmercury. In contrast, the dopamine induced response was not affected by methylmercury at 30 microM. It was concluded that methylmercury impairs both sodium and potassium channel gating mechanisms and suppresses acetylcholine receptor/channel complexes. It remains to be seen whether the effect of chronic exposure is similar to that seen after acute and high level exposure in the present study.     

Biophysical properties of scorpion alpha-toxin-sensitive background sodium channel contributing to the pacemaker activity in insect neurosecretory cells (DUM neurons)

A scorpion alpha-toxin-sensitive background sodium channel was characterized in short-term cultured adult cockroach dorsal unpaired median (DUM) neurons using the cell-attached patch-clamp configuration. Under control conditions, spontaneous sodium currents were recorded at different steady-state holding potentials, including the range of normal resting membrane potential. At -50 mV, the sodium current was observed as unclustered, single openings. For potentials more negative than -70 mV, investigated patches contained large unitary current steps appearing generally in bursts. These background channels were blocked by tetrodotoxin (TTX, 100 nm), and replacing sodium with TMA-Cl led to a complete loss of channel activity. The current-voltage relationship has a slope conductance of 36 pS. At -50 mV, the mean open time constant was 0.22 +/- 0.05 ms (n = 5). The curve of the open probability versus holding potentials was bell-shaped, with its maximum (0.008 +/- 0.004; n = 5) at -50 mV. LqhalphaIT (10-8 m) altered the background channel activity in a time-dependent manner. At -50 mV, the channel activity appeared in bursts. The linear current-voltage relationship of the LqhalphaIT-modified sodium current determined for the first three well-resolved open states gave three conductance levels: 34, 69 and 104 pS, and reversed at the same extrapolated reversal potential (+52 mV). LqhalphaIT increased the open probability but did not affect either the bell-shaped voltage dependence or the open time constant. Mammal toxin AaHII induced very similar effects on background sodium channels but at a concentration 100 x higher than LqhalphaIT. At 10-7 m, LqhalphaIT produced longer silence periods interrupted by bursts of increased channel activity. Whole-cell experiments suggested that background sodium channels can provide the depolarizing drive for DUM neurons essential to maintain beating pacemaker activity, and revealed that 10-7 m LqhalphaIT transformed a beating pacemaker activity into a rhythmic bursting.