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The ionic mechanisms underlying the termination of action-potential (AP) bursts and postburst afterhyperpolarization (AHP) in intrinsically bursting (IB) neocortical neurons were investigated by performing intracellular recordings in thin slices of rat sensorimotor cortex. The blockade of Ca(2+)-activated K(+) currents enhanced postburst depolarizing afterpotentials, but had inconsistent and minor effects on the amplitude and duration of AHPs. On the contrary, experimental conditions resulting in reduction of voltage-dependent Na(+) entry into the cells caused a significant decrease of AHP amplitude. Slice perfusion with a modified artificial cerebrospinal fluid in which LiCl (40 mM) partially replaced NaCl had negligible effects on the properties of individual APs, whereas it consistently increased burst length and led to an approximately 30% reduction in the amplitude of AHPs following individual bursts or short trains of stimulus-induced APs. Experiments performed by partially replacing Na(+) ions with choline revealed a comparable reduction in AHP amplitude associated with an inhibition of bursting activity. Moreover, in voltage-clamp experiments carried out in both in situ and acutely isolated neurons, partial substitution of extracellular NaCl with LiCl significantly and reversibly reduced the amplitude of K(+) currents evoked by depolarizing stimuli above-threshold for Na(+)-current activation. The above effect of Na(+)-to-Li(+) substitution was not seen when voltage-gated Na(+) currents were blocked with TTX, indicating the presence of a specific K(+)-current component activated by voltage-dependent Na(+) (but not Li(+)) influx. The above findings suggest that a Na(+)-activated K(+) current recruited by the Na(+) entry secondary to burst discharge significantly contributes to AHP generation and the maintenance of rhythmic burst recurrence during sustained depolarizations in neocortical IB neurons.

Knockout Otx1 mice present a microcephalic phenotype mainly due to reduced deep neocortical layers and spontaneous recurrent seizures. We investigated the excitable properties of layer V pyramidal neurons in neocortical slices prepared from Otx1-/- mice and age-matched controls. The qualitative firing properties of the neurons of Otx1-/- mice were identical to those found in wild-type controls, but the proportion of intrinsically bursting (IB) neurons was significantly smaller. This is in line with the lack of the Otx1 gene contribution to the generation and differentiation of neurons destined for the deep neocortical layers, in which IB neurons are located selectively in wild-type rodents. The pyramidal neurons recorded in Otx1-/- mice responded to near-threshold electrical stimulation of the underlying white matter, with aberrant polysynaptic excitatory potentials often leading to late action potential generation. When the strength of the stimulus was increased, the great majority of the Otx1-/- neurons (78%) responded with a prominent biphasic inhibitory postsynaptic potential that was significantly larger than that observed in the wild-type mice, and was often followed by complex postinhibitory depolarizing events. Both late excitatory postsynaptic potentials and postinhibitory excitation were selectively suppressed by NMDA receptor antagonists, but not by AMPA antagonists. We conclude that the cortical abnormalities of Otx1-/- neocortex due to a selective loss of large projecting neurons lead to a complex rearrangement of local circuitry, which is characterized by an excess of N-methyl-D-aspartate-mediated polysynaptic excitation that is counteracted by GABA-mediated inhibition in only a limited range of stimulus intensity. Prominent postsynaptic inhibitory potentials may also act as a further pro-epileptogenic event by synchronizing abnormal excitatory potentials.

The murine homeobox-containing Otx gene is required for correct nervous system and sense organ development. Otx1-/1 mice obtained by replacing Otx with the lac Z gene show developmental abnormalities of the cerebellum, mesencephalon, and cerebral cortex associated with spontaneous epileptic seizures (1). The epileptogenic mechanisms accounting for these seizures were investigated by means of electrophysiological recordings made from neocortical slices.

The effect of the protein kinase C (PKC) activator 1-oleoyl-2-acetyl-sn-glycerol (OAG) on TTX-sensitive Na+ currents in neocortical pyramidal neurones was evaluated using voltage-clamp and intracellular current-clamp recordings. In pyramid-shaped dissociated neurones, the addition of OAG to the superfusing medium consistently led to a 30% reduction in the maximal peak amplitude of the transient sodium current (I(Na,T)) evoked from a holding potential of -70 mV. We attributed this inhibitory effect to a significant negative shift of the voltage dependence of steady-state channel inactivation (of approximately 14 mV). The inhibitory effect was completely prevented by hyperpolarising prepulses to potentials that were more negative than -80 mV. A small but significant leftward shift of INa,T activation was also observed, resulting in a slight increase of the currents evoked by test pulses at potentials more negative then -35 mV. In the presence of OAG, the activation of the persistent fraction of the Na+ current (INa,P) evoked by means of slow ramp depolarisations was consistently shifted in the negative direction by 3.9+/-0.5 mV, while the peak amplitude of the current was unaffected. In slice experiments, the OAG perfusion enhanced a subthreshold depolarising rectification affecting the membrane response to the injection of positive current pulses, and thus led the neurones to fire in response to significantly lower depolarising stimuli than those needed under control conditions. This effect was attributed to an OAG-induced enhancement of INa,P, since it was observed in the same range of potentials over which I(Na,P) activates and was completely abolished by TTX. The qualitative firing characteristics of both the intrinsically bursting and regular spiking neurones were unaffected when OAG was added to the physiological perfusing medium, but their firing frequency increased in response to slight suprathreshold depolarisations. The obtained results suggest that physiopa

The maturational profile of the firing characteristics of 217 layer V pyramidal neurons of rat sensorimotor cortex, injected with biocytin for morphological reconstruction, was analysed by means of intracellular recordings made between postnatal day (P)3 and 22. Starting from the onset of the second postnatal week, the pyramidal neurons could be differentiated as adapting or non-adapting regular spiking on the basis of the presence or absence of spike frequency adaptation. The percentage of non-adapting regular spiking neurons was very high during the second postnatal week (53%) and progressively decreased with age, concurrently with the appearance of the new class of intrinsically bursting neurons (beginning of the third week) whose percentage progressively increased from 23%, found in P14-P16 rats, to 46% in adult rats. Non-adapting regular spiking neurons were found to share with intrinsically bursting neurons several physiological characteristics comprehending faster action potentials, more prominent effect of anomalous rectification and consistent depolarizing afterpotentials, that differentiated them from the adapting regular spiking neurons. Moreover, intrinsically bursting and non-adapting regular spiking neurons were characterized by a round-shaped distribution of basal dendrites and expanded apical dendritic arborization, that differentiated them from the adapting regular spiking neurons showing a simpler dendritic arborization. These morphological hallmarks were seen in immature intrinsically bursting neurons as soon as they became distinguishable, and in immature non-adapting regular spiking neurons starting from the onset of the second postnatal week. These findings suggest that a significant subpopulation of immature non-adapting regular spiking neurons are committed to becoming bursters, and that they are converted into intrinsically bursting neurons during the second postnatal week, as soon as the ionic current sustaining the burst firing is sufficie

In in vitro slices prepared from rat sensorimotor cortex, intracellular recordings were obtained from 107 layer V pyramidal neurons, subsequently injected with biocytin for morphological reconstruction. Of the 107 neurons, 59 (55.1%) were identified as adapting (45) or non-adapting (13) regular spiking neurons (RS), and 48 (44.9%) as intrinsically bursting (IB) neurons discharging with an initial cluster of action potentials, which tended to recur rhythmically in a subset of 19 cells. The block of IAR by extracellular Cs+ did not affect burst generation, but enhanced the tendency to reburst in IB neurons. A similar effect was induced by other procedures affecting K(+)-dependent post-burst hyperpolarization. In IB neurons Ca2+ spikes had a longer decay time than in RS neurons, however selective blockers of both low and high threshold Ca2+ conductances failed to impair bursting activity. On the contrary, the perfusion of the slices with 0.5-1 microM TTX suppressed bursting behaviour in a critical time interval preceding the complete block of Na(+)-dependent action potentials. It is concluded that the persistent Na+ current INAP is the most important intrinsic factor for the typical firing properties of IB neurons, while Ca2+ and K+ conductances appear to contribute towards shaping bursts and controlling their recurrence rate. The morphology, connectivity and physiological properties of adapting and non-adapting RS neurons are particularly suited to the processing of respectively phasic and tonic inputs, whereas the properties of IB neurons are consistent with their suggested role in cortical rhythmogenesis and in the pathophysiological synchronized activities underlying epileptogenesis.

Intrinsically bursting (IB) neurons, responding with a burst of action potentials to just threshold intracellular depolarizing current pulses, are encountered in layer V of mature rodent sensorimotor cortex. We report the results of intracellular recordings performed on neocortical slices obtained from immature rats between postnatal day (P) 7 and P21, as compared to adult animals (above P60). The bursting properties are here reported to mature abruptly around P14. After this time a subpopulation of IB neurons was recognizable on the basis of both physiological and morphological characteristics (i.e. extensive apical and basal dendrites arborization, axon collaterals limited to layers V-VI). Maturational changes in number and distribution of Ca2+/K+ channels may account for this developmental step. The immaturity of IB neurons may be correlated with the poorly synchronized character of cortical activities in the very young animals.