Contributions of Kv7-mediated potassium current to sub- and suprathreshold responses of rat layer II/III neocortical pyramidal neurons.

After block of Kv1- and Kv2-mediated K(+) currents in acutely dissociated neocortical pyramidal neurons from layers II/III of rat somatosensory and motor cortex, the remaining current is slowly activating and persistent. We used whole cell voltage clamp to show that the Kv7 blockers linopirdine and XE-991 blocked a current with similar kinetics to the current remaining after combined block of Kv1 and Kv2 channels. This current was sensitive to low doses of linopirdine and activated more slowly and at more negative potentials than Kv1- or Kv2-mediated current. The Kv7-mediated current decreased in amplitude with time in whole cell recordings, but in most cells the current was stable for several minutes. Current in response to a traditional M-current protocol was blocked by muscarine, linopirdine, and XE-991. Whole cell slice recordings revealed that the Q10 for channel deactivation was ∼2.5. Sharp electrode current-clamp recordings from adult pyramidal cells demonstrated that block of Kv7-mediated current with XE-991 reduced rheobase, shortened the latency to firing to near rheobase current, induced more regular firing at low current intensity, and increased the rate of firing to a given current injection. XE-991 did not affect single action potentials or spike frequency adaptation. Application of XE-991 also eliminated subthreshold voltage oscillations and increased gain for low-frequency inputs (<10 Hz) without affecting gain for higher frequency inputs. These data suggest important roles for Kv7 channels in subthreshold regulation of excitability, generation of theta-frequency subthreshold oscillations, regulation of interspike intervals, and biasing selectivity toward higher frequency inputs.

Endogenous calcium buffering capacity of substantia nigral dopamine neurons.

Dopamine (DA)-containing cells from the substantia nigra pars compacta (SNc) play a major role in the initiation of movement. Loss of these cells results in Parkinson's disease (PD). Changes in intracellular calcium ion concentration ([Ca(2+)](i)) elicit several events in DA cells, including spike afterhyperpolarizations (AHPs) and subthreshold oscillations underlying autonomous firing. Continuous Ca(2+) load due to Ca(2+)-dependent rhythmicity has been proposed to cause the death of DA cells in PD and normal aging. Because of the physiological and pathophysiological importance of [Ca(2+)](i) in DA cells, we characterized their intrinsic Ca(2+)-buffering capacity (K(S)) in brain slices. We introduced a fluorescent Ca(2+)-sensitive exogenous buffer (200 microM fura-2) and cells were tracked from break-in until steady state by stimulating with a single action potential (AP) every 30 s and measuring the Ca(2+) transient from the proximal dendrite. DA neurons filled exponentially with a tau of about 5-6 min. [Ca(2+)](i) was assumed to equilibrate between the endogenous Ca(2+) buffer and the exogenous Ca(2+) indicator buffer. Intrinsic buffering was estimated by extrapolating from the linear relationships between the amplitude or time constant of the Ca(2+) transients versus [fura-2]. Extrapolated Ca(2+)-transients in the absence of fura-2 had mean peak amplitudes of 293.7 +/- 65.3 nM and tau = 124 +/- 13 ms (postnatal day 13 [P13] to P17 animals). Intrinsic buffering increased with age in DA neurons. For cells from animals P13-P17, K(S) was estimated to be about 110 (n = 20). In older animals (P25-P32), the estimate was about 179 (n = 10). These relatively low values may reflect the need for rapid Ca(2+) signaling, e.g., to allow activation of sK channels, which shape autonomous oscillations and burst firing. Low intrinsic buffering may also make DA cells vulnerable to Ca(2+)-dependent pathology.

Kv2 subunits underlie slowly inactivating potassium current in rat neocortical pyramidal neurons.

We determined the expression of Kv2 channel subunits in rat somatosensory and motor cortex and tested for the contributions of Kv2 subunits to slowly inactivating K+ currents in supragranular pyramidal neurons. Single cell RT-PCR showed that virtually all pyramidal cells expressed Kv2.1 mRNA and approximately 80% expressed Kv2.2 mRNA. Immunocytochemistry revealed striking differences in the distribution of Kv2.1 and Kv2.2 subunits. Kv2.1 subunits were clustered and located on somata and proximal dendrites of all pyramidal cells. Kv2.2 subunits were primarily distributed on large apical dendrites of a subset of pyramidal cells from deep layers. We used two methods for isolating currents through Kv2 channels after excluding contributions from Kv1 subunits: intracellular diffusion of Kv2.1 antibodies through the recording pipette and extracellular application of rStromatoxin-1 (ScTx). The Kv2.1 antibody specifically blocked the slowly inactivating K+ current by 25-50% (at 8 min), demonstrating that Kv2.1 subunits underlie much of this current in neocortical pyramidal neurons. ScTx (300 nM) also inhibited approximately 40% of the slowly inactivating K+ current. We observed occlusion between the actions of Kv2.1 antibody and ScTx. In addition, Kv2.1 antibody- and ScTx-sensitive currents demonstrated similar recovery from inactivation and voltage dependence and kinetics of activation and inactivation. These data indicate that both agents targeted the same channels. Considering the localization of Kv2.1 and 2.2 subunits, currents from truncated dissociated cells are probably dominated by Kv2.1 subunits. Compared with Kv2.1 currents in expression systems, the Kv2.1 current in neocortical pyramidal cells activated and inactivated at relatively negative potentials and was very sensitive to holding potential.

Functional roles of Kv1 channels in neocortical pyramidal neurons.

Pyramidal neurons from layers II/III of somatosensory and motor cortex express multiple Kv1 alpha-subunits and a current sensitive to block by alpha-dendrotoxin (alpha-DTX). We examined functional roles of native Kv1 channels in these cells using current-clamp recordings in brain slices and current- and voltage-clamp recordings in dissociated cells. alpha-DTX caused a significant negative shift in voltage threshold for action potentials (APs) and reduced rheobase. Correspondingly, a ramp-voltage protocol revealed that the alpha-DTX-sensitive current activated at subthreshold voltages. AP width at threshold increased with successive APs during repetitive firing. The steady-state threshold width for a given firing rate was similar in control and alpha-DTX, despite an initially broader AP in alpha-DTX. AP voltage threshold increased similarly during a train of spikes under control conditions and in the presence of alpha-DTX. alpha-DTX had no effect on input resistance or resting membrane potential and modest effects on the amplitude or width of a single AP. Accordingly, experiments using AP waveforms (APWs) as voltage protocols revealed that alpha-DTX-sensitive current peaked late during the AP repolarization phase. Application of alpha-DTX increased the rate of firing to intracellular current injection and increased gain (multiplicative effects), but did not alter spike-frequency adaptation. Consistent with these findings, voltage-clamp experiments revealed that the proportion of outward current sensitive to alpha-DTX was highest during the interval between two APWs, reflecting slow deactivation kinetics at -50 mV. Finally, alpha-DTX did not alter the selectivity of pyramidal neurons for DC versus time-varying stimuli.

Expression and biophysical properties of Kv1 channels in supragranular neocortical pyramidal neurones.

Potassium channels are extremely diverse regulators of neuronal excitability. As part of an investigation into how this molecular diversity is utilized by neurones, we examined the expression and biophysical properties of native Kv1 channels in layer II/III pyramidal neurones from somatosensory and motor cortex. Single-cell RT-PCR, immunocytochemistry, and whole cell recordings with specific peptide toxins revealed that individual pyramidal cells express multiple Kv1 alpha-subunits. The most abundant subunit mRNAs were Kv1.1 > 1.2 > 1.4 > 1.3. All of these subunits were localized to somatodendritic as well as axonal cell compartments. These data suggest variability in the subunit complexion of Kv1 channels in these cells. The alpha-dendrotoxin (alpha-DTX)-sensitive current activated more rapidly and at more negative potentials than the alpha-DTX-insensitive current, was first observed at voltages near action potential threshold, and was relatively insensitive to holding potential. The alpha-DTX-sensitive current comprised about 10% of outward current at steady-state, in response to steps from -70 mV. From -50 mV, this percentage increased to approximately 20%. All cells expressed an alpha-DTX-sensitive current with slow inactivation kinetics. In some cells a transient component was also present. Deactivation kinetics were voltage dependent, such that deactivation was slow at potentials traversed by interspike intervals during repetitive firing. Because of its kinetics and voltage dependence, the alpha-DTX-sensitive current should be most important at physiological resting potentials and in response to brief stimuli. Kv1 channels should also be important at voltages near threshold and corresponding to interspike intervals.

Effects of temperature on calcium transients and Ca2+-dependent afterhyperpolarizations in neocortical pyramidal neurons.

In neocortical pyramidal neurons, the medium (mAHP) and slow AHP (sAHP) have different relationships with intracellular [Ca2+]. To further explore these differences, we varied bath temperature and compared passive and active membrane properties and Ca2+ transients in response to a single action potential (AP) or trains of APs. We tested whether Ca(2+)-dependent events are more temperature sensitive than voltage-dependent ones, the slow rise time of the sAHP is limited by diffusion, and temperature sensitivity differs between the mAHP and sAHP. The onset and decay kinetics of the sAHP were very temperature sensitive (more so than diffusion). We found that the decay time course of Ca2+ transients was also very temperature sensitive. In contrast, the mAHP (amplitude, time to peak, and exponential decay) and sAHP peak amplitude were moderately sensitive to temperature. The amplitudes of intracellular Ca2+ transients evoked either by a single spike or a train of spikes showed modest temperature sensitivities. Pyramidal neuron input resistance was increased by cooling. With the exception of threshold, which remained unchanged between 22 and 35 degrees C, action potential parameters (amplitude, half-width, maximum rates of rise and fall) were modestly affected by temperature. Collectively, these data suggest that temperature sensitivity was higher for the Ca(2+)-dependent sAHP than for voltage-dependent AP parameters or for the mAHP, diffusion of Ca2+ over distance cannot explain the slow rise of the sAHP in these cells, and the kinetics of the sAHP and mAHP are affected differently by temperature.

alpha2-Adrenergic receptor-mediated modulation of calcium current in neocortical pyramidal neurons.

Noradrenergic projections to the cortex modulate a variety of cortical activities and calcium channels are one likely target for such modulation. We used the whole-cell patch-clamp technique to study noradrenergic modulation of barium currents in acutely dissociated pyramidal neurons from rat sensorimotor cortex. Extracellular application of specific agonists and antagonists revealed that norepinephrine (NE) reduced Ca2+ current. A major component of this modulation was due to activation of alpha2 receptors. Activation of alpha2-adrenergic receptors resulted in a fast, voltage-dependent pathway involving Gi/Go G-proteins. This pathway targeted N- and P-type calcium channels The alpha2 modulation was partially reversed by repeated action potential waveforms (APWs). N- and P-type channels have been implicated in synaptic transmission and activation of afterhyperpolarizations in these cells. Our findings suggest that NE can regulate these cellular processes by mechanisms sensitive to spike activity.

Relationships between intracellular calcium and afterhyperpolarizations in neocortical pyramidal neurons.

We examined the effects of recent discharge activity on [Ca2+]i in neocortical pyramidal cells. Our data confirm and extend the observation that there is a linear relationship between plateau [Ca2+]i and firing frequency in soma and proximal apical dendrites. The rise in [Ca2+] activates K+ channels underlying the afterhyperpolarization (AHP), which consists of 2 Ca(2+)-dependent components: the medium AHP (mAHP) and the slow AHP (sAHP). The mAHP is blocked by apamin, indicating involvement of SK-type Ca(2+)-dependent K+ channels. The identity of the apamin-insensitive sAHP channel is unknown. We compared the sAHP and the mAHP with regard to: 1) number and frequency of spikes versus AHP amplitude; 2) number and frequency of spikes versus [Ca2+]i; 3) IAHP versus [Ca2+]i. Our data suggest that sAHP channels require an elevation of [Ca2+]i in the cytoplasm, rather than at the membrane, consistent with a role for a cytoplasmic intermediate between Ca2+ and the K+ channels. The mAHP channels appear to respond to a restricted Ca2+ domain.

Serotonergic modulation of supragranular neurons in rat sensorimotor cortex.

Numerous observations suggest diverse and modulatory roles for serotonin (5-HT) in cortex. Because of the diversity of cell types and multiple receptor subtypes and actions of 5-HT, it has proven difficult to determine the overall role of 5-HT in cortical function. To provide a broader perspective of cellular actions, we studied the effects of 5-HT on morphologically and physiologically identified pyramidal and nonpyramidal neurons from layers I-III of primary somatosensory and motor cortex. We found cell type-specific differences in response to 5-HT. Four cell types were observed in layer I: Cajal Retzius, pia surface, vertical axon, and horizontal axon cells. The physiology of these cells ranged from fast spiking (FS) to regular spiking (RS). In layers II-III, we observed interneurons with FS, RS, and late spiking physiology. Morphologically, these cells varied from bipolar to multipolar and included basket-like and chandelier cells. 5-HT depolarized or hyperpolarized pyramidal neurons and reduced the slow afterhyperpolarization and spike frequency. Consistent with a role in facilitating tonic inhibition, 5-HT2 receptor activation increased the frequency of spontaneous IPSCs in pyramidal neurons. In layers II-III, 70% of interneurons were depolarized by 5-HT. In layer I, 57% of cells with axonal projections to layers II-III (vertical axon) were depolarized by 5-HT, whereas 63% of cells whose axons remain in layer I (horizontal axon) were hyperpolarized by 5-HT. We propose a functional segregation of 5-HT effects on cortical information processing, based on the pattern of axonal arborization.

Effects of spike parameters and neuromodulators on action potential waveform-induced calcium entry into pyramidal neurons.

Neocortical pyramidal neurons express several different calcium channel types. Previous studies with square voltage steps have found modest biophysical differences between these calcium channel types as well as differences in their modulation by transmitters. We used acutely dissociated neocortical pyramidal neurons to test whether this diversity extends to different activation by physiological stimuli. We conclude that 1) peak amplitude, latency to peak, and the total charge entry for the Ca(2+) channel current is dependent on the shape of the mock action potential waveforms (APWs). 2) The percent contribution of the five high-voltage-activated currents to the whole cell current was not altered by using an APW as opposed to a voltage step to elicit the current. 3) The identity of the charge carrier affects the amplitude and decay of the whole cell current. With Ca(2+), there was a greater contribution of T-type current to the whole cell current. 4) Total Ba(2+) charge entry is linearly dependent on the number of spikes in the stimulating waveform and relatively insensitive to spike frequency. 5) Current decay was greatest with Ca(2+) as the charge carrier and with minimal internal chelation. 6) Voltage-dependent neurotransmitter-mediated modulations can be reversed by multiple spikes. The extent of the reversal is dependent on the number of spikes in the stimulating waveform. Thus the neuronal activity pattern can determine the effectiveness of voltage-dependent and -independent modulatory pathways in neocortical pyramidal neurons.

Morphological and electrophysiological properties of atypically oriented layer 2 pyramidal cells of the juvenile rat neocortex.

We used whole-cell patch clamp recordings combined with intracellular dye-filling to examine the morphological and electrophysiological properties of atypically oriented pyramidal cells located at the layer 1/2 border of the juvenile rat neocortex. Orientation of the apical dendrite varied from oblique (>20 degrees from vertical) to truly horizontal (90 degrees from vertical). The length of the apical dendrite ranged from 150 to 400 microm. The total horizontal domain of the dendritic tree (including basal dendrites) of the longest horizontal pyramids exceeded 500 microm, but we also found short horizontal cells with horizontal dendritic domains of less than 300 microm. In addition, atypically oriented pyramids had long horizontal axon collaterals in layer 1/2. Electrophysiologically, atypically oriented pyramidal cells had intrinsic membrane properties similar to regularly oriented pyramids that have been described in the superficial layers at this age in the rat. Cells that fired repetitively were all regular spiking. In addition, we identified a subgroup of neurons (20%) in this sample, which were unable to fire more than a few spikes at the beginning of the current pulse. We suggest that the unique orientation and size of their dendritic trees and the length and arrangement of their local axons collaterals make atypically oriented pyramids in layer 2 ideally suited to perform horizontal integration of synaptic inputs in the neocortex.

Unique properties of R-type calcium currents in neocortical and neostriatal neurons.

Whole cell recordings from acutely dissociated neocortical pyramidal neurons and striatal medium spiny neurons exhibited a calcium-channel current resistant to known blockers of L-, N-, and P/Q-type Ca(2+) channels. These R-type currents were characterized as high-voltage-activated (HVA) by their rapid deactivation kinetics, half-activation and half-inactivation voltages, and sensitivity to depolarized holding potentials. In both cell types, the R-type current activated at potentials relatively negative to other HVA currents in the same cell type and inactivated rapidly compared with the other HVA currents. The main difference between cell types was that R-type currents in neocortical pyramidal neurons inactivated at more negative potentials than R-type currents in medium spiny neurons. Ni(2+) sensitivity was not diagnostic for R-type currents in either cell type. Single-cell RT-PCR revealed that both cell types expressed the alpha1E mRNA, consistent with this subunit being associated with the R-type current.

Calcium currents in retrogradely labeled pyramidal cells from rat sensorimotor cortex.

Our previous studies of calcium (Ca(2+)) currents in cortical pyramidal cells revealed that the percentage contribution of each Ca(2+) current type to the whole cell Ca(2+) current varies from cell to cell. The extent to which these currents are modulated by neurotransmitters is also variable. This study was directed at testing the hypothesis that a major source of this variability is recording from multiple populations of pyramidal cells. We used the whole cell patch-clamp technique to record from dissociated corticocortical, corticostriatal, and corticotectal projecting pyramidal cells. There were significant differences between the three pyramidal cell types in the mean percentage of L-, P-, and N-type Ca(2+) currents. For both N- and P-type currents, the range of percentages expressed was small for corticostriatal and corticotectal cells as compared with cells which project to the corpus callosum or to the general population. The variance was significantly different between cell types for N- and P-type currents. These results suggest that an important source of the variability in the proportions of Ca(2+) current types present in neocortical pyramidal neurons is recording from multiple populations of pyramidal cells.

Different Ca2+ source for slow AHP in completely adapting and repetitive firing pyramidal neurons.

Intracellular recordings in an in vitro neocortical slice preparation from immature rats were used to investigate the Ca2 source for slow afterhyperpolarization (sAHP) generation in pyramidal neurons that exhibit complete spike frequency adaptation (CA neurons). In pyramidal neurons that maintain repetitive firing for long periods of time (RF neurons), N-, P- and Q-type Ca2+ channels supply Ca2+ for sAHP generation. In CA neurons, the sAHP was reduced by only 50% by the combination of antagonists for these Ca2+ channel types and L-type channels. Ryanodine and dantrolene, blockers of Ca2(+)-induced Ca2+ release, reduced the sAHP by approximately 45% in CA neurons, but caused no reduction of the sAHP in RF neurons. Dantrolene application caused CA neurons to fire throughout a 1s suprathreshold current injection (as do RF neurons).

Properties of Q-type calcium channels in neostriatal and cortical neurons are correlated with beta subunit expression.

In brain neurons, P- and Q-type Ca(2+) channels both appear to include a class A alpha1 subunit. In spite of this similarity, these channels differ pharmacologically and biophysically, particularly in inactivation kinetics. The molecular basis for this difference is unclear. In heterologous systems, alternative splicing and ancillary beta subunits have been shown to alter biophysical properties of channels containing a class A alpha1 subunit. To test the hypothesis that similar mechanisms are at work in native systems, P- and Q-type currents were characterized in acutely isolated rat neostriatal, medium spiny neurons and cortical pyramidal neurons using whole-cell voltage-clamp techniques. Cells were subsequently aspirated and subjected to single-cell RT-PCR (scRT-PCR) analysis of calcium channel alpha(1) and beta (beta(1-4)) subunit expression. In both cortical and neostriatal neurons, P- and Q-type currents were found in cells expressing class A alpha(1) subunit mRNA. Although P-type currents in cortical and neostriatal neurons were similar, Q-type currents differed significantly in inactivation kinetics. Notably, Q-type currents in neostriatal neurons were similar to P-type currents in inactivation rate. The variation in Q-type channel biophysics was correlated with beta subunit expression. Neostriatal neurons expressed significantly higher levels of beta(2a) mRNA and lower levels of beta(1b) mRNA than cortical neurons. These findings are consistent with the association of beta(2a) and beta(1b) subunits with slow and fast inactivation, respectively. Analysis of alpha(1A) splice variants in the linker between domains I and II failed to provide an alternative explanation for the differences in inactivation rates. These findings are consistent with the hypothesis that the biophysical properties of Q-type channels are governed by beta subunit isoforms and are separable from toxin sensitivity.

Neuromodulation, development and synaptic plasticity.

We discuss parallels in the mechanisms underlying use-dependent synaptic plasticity during development and long-term potentiation (LTP) and long-term depression (LTD) in neocortical synapses. Neuromodulators, such as norepinephrine, serotonin, and acetylcholine have also been implicated in regulating both developmental plasticity and LTP/LTD. There are many potential levels of interaction between neuromodulators and plasticity. Ion channels are substrates for modulation in many cell types. We discuss examples of modulation of voltage-gated Ca2+ channels and Ca(2+)-dependent K+ channels and the consequences for neocortical pyramidal cell firing behaviour. At the time when developmental plasticity is most evident in rat cortex, the substrate for modulation is changing as the densities and relative proportions of various ion channels types are altered during ontogeny. We discuss examples of changes in K+ and Ca2+ channels and the consequence for modulation of neuronal activity.

Differences in the properties of ionotropic glutamate synaptic currents in oxytocin and vasopressin neuroendocrine neurons.

Oxytocin (OT) and vasopressin (VP) hormone release from neurohypophysial terminals is controlled by the firing pattern of neurosecretory cells located in the hypothalamic supraoptic (SON) and paraventricular nuclei. Although glutamate is a key modulator of the electrical activity of both OT and VP neurons, a differential contribution of AMPA receptors (AMPARs) and NMDA receptors (NMDARs) has been proposed to mediate glutamatergic influences on these neurons. In the present study we examined the distribution and functional properties of synaptic currents mediated by AMPARs and NMDARs in immunoidentified SON neurons. Our results suggest that the properties of AMPA-mediated currents in SON neurons are controlled in a cell type-specific manner. OT neurons displayed AMPA-mediated miniature EPSCs (mEPSCs) with larger amplitude and faster decay kinetics than VP neurons. Furthermore, a peak-scaled nonstationary noise analysis of mEPSCs revealed a larger estimated single-channel conductance of AMPARs expressed in OT neurons. High-frequency summation of AMPA-mediated excitatory postsynaptic potentials was smaller in OT neurons. In both cell types, AMPA-mediated synaptic currents showed inward rectification, which was more pronounced in OT neurons, and displayed Ca2+ permeability. On the other hand, NMDA-mediated mEPSCs of both cell types had similar amplitude and kinetic properties. The cell type-specific expression of functionally different AMPARs can contribute to the adoption of different firing patterns by these neuroendocrine neurons in response to physiological stimuli.

Muscarine modulates Ca2+ channel currents in rat sensorimotor pyramidal cells via two distinct pathways.

We used the whole cell patch-clamp technique and single-cell reverse transcription-polymerase chain reaction (RT-PCR) to study the muscarinic receptor-mediated modulation of calcium channel currents in both acutely isolated and cultured pyramidal neurons from rat sensorimotor cortex. Single-cell RT-PCR profiling for muscarinic receptor mRNAs revealed the expression of m1, m2, m3, and m4 subtypes in these cells. Muscarine reversibly reduced Ca2+ currents in a dose-dependent manner. The modulation was blocked by the muscarinic antagonist atropine. When the internal recording solution included 10 mM ethylene glycol-bis(beta-aminoethyl ether)-N, N,N',N'-tetraacetic acid (EGTA) or 10 mM bis-(o-aminophenoxy)-N,N,N', N'-tetraacetic acid (BAPTA), the modulation was rapid (tauonset approximately 1.2 s). Under conditions where intracellular calcium levels were less controlled (0.0-0.1 mM BAPTA), a slowly developing component of the modulation also was observed (tauonset approximately 17 s). Both fast and slow components also were observed in recordings with 10 mM EGTA or 20 mM BAPTA when Ca2+ was added to elevate internal [Ca2+] ( approximately 150 nM). The fast component was due to a reduction in both N- and P-type calcium currents, whereas the slow component involved L-type current. N-ethylmaleimide blocked the fast component but not the slow component of the modulation. Preincubation of cultured neurons with pertussis toxin (PTX) also greatly reduced the fast portion of the modulation. These results suggest a role for both PTX-sensitive G proteins as well as PTX-insensitive G proteins in the muscarinic modulation. The fast component of the modulation was reversed by strong depolarization, whereas the slow component was not. Reblock of the calcium channels by G proteins (at -90 mV) occurred with a median tau of 68 ms. We conclude that activation of muscarinic receptors results in modulation of N- and P-type channels by a rapid, voltage-dependent pathway and of L-type current by a slow, voltage-independent pathway.

Specificity in the interaction of HVA Ca2+ channel types with Ca2+-dependent AHPs and firing behavior in neocortical pyramidal neurons.

Intracellular recordings and organic and inorganic Ca2+ channel blockers were used in a neocortical brain slice preparation to test whether high-voltage-activated (HVA) Ca2+ channels are differentially coupled to Ca2+-dependent afterhyperpolarizations (AHPs) in sensorimotor neocortical pyramidal neurons. For the most part, spike repolarization was not Ca2+ dependent in these cells, although the final phase of repolarization (after the fast AHP) was sensitive to block of N-type current. Between 30 and 60% of the medium afterhyperpolarization (mAHP) and between approximately 80 and 90% of the slow AHP (sAHP) were Ca2+ dependent. Based on the effects of specific organic Ca2+ channel blockers (dihydropyridines, omega-conotoxin GVIA, omega-agatoxin IVA, and omega-conotoxin MVIIC), the sAHP is coupled to N-, P-, and Q-type currents. P-type currents were coupled to the mAHP. L-type current was not involved in the generation of either AHP but (with other HVA currents) contributes to the inward currents that regulate interspike intervals during repetitive firing. These data suggest different functional consequences for modulation of Ca2+ current subtypes.

Fast-to-slow conversion following chronic low-frequency activation of medial gastrocnemius muscle in cats. II. Motoneuron properties.

Chronic stimulation (for 2-3 mo) of the medial gastrocnemius (MG) muscle nerve by indwelling electrodes renders the normally heterogeneous MG muscle mechanically and histochemically slow (type SO). We tested the hypothesis that motoneurons of MG muscle thus made type SO by chronic stimulation would also convert to slow phenotype. Properties of all single muscle units became homogeneously type SO (slowly contracting, nonfatiguing, nonsagging contraction during tetanic activation). Motoneuron electrical properties were also modified in the direction of type S, fatigue-resistant motor units. Two separate populations were identified (on the basis of afterhyperpolarization, rheobase, and input resistance) that likely correspond to motoneurons that had been fast (type F) or type S before stimulation. Type F motoneurons, although modified by chronic stimulation, were not converted to the type S phenotype, despite apparent complete conversion of their muscle units to the slow oxidative type (type SO). Muscle units of the former type F motor units were faster and/or more powerful than those of the former type S motor units, indicating some intrinsic regulation of motor unit properties. Experiments in which chronic stimulation was applied to the MG nerve cross-regenerated into skin yielded changes in motoneuron properties similar to those above, suggesting that muscle was not essential for the effects observed. Modulation of group Ia excitatory postsynaptic potential (EPSP) amplitude during high-frequency trains, which in normal MG motoneurons can be either positive or negative, was negative in 48 of 49 chronically stimulated motoneurons. Negative modulation is characteristic of EPSPs in motoneurons of most fatigue-resistant motor units. The general hypothesis of a periphery-to-motoneuron retrograde mechanism was supported, although the degree of control exerted by the periphery may vary: natural type SO muscle appears especially competent to modify motoneuron properties. We speculate that activity-dependent regulation of the neurotrophin-(NT) 4/5 in muscle plays an important role in controlling muscle and motoneuron properties.