Slow adaptation in fast-spiking neurons of visual cortex.

Fast-spiking (FS) neurons are a class of inhibitory interneurons classically characterized as having short-duration action potentials (<0.5 ms at half height) and displaying little to no spike-frequency adaptation during short (<500 ms) depolarizing current pulses. As a consequence, the resulting injected current intensity versus firing frequency relationship is typically steep, and they can achieve firing frequencies of < or =1 kHz. Here we have investigated the properties of FS neurons discharges on a longer time scale. Twenty second discharges were induced in electrophysiologically identified FS neurons by means of current injection either with sinusoidal current or with square pulses. We found that virtually all FS neurons recorded in cortical slices do show spike-frequency adaptation but with a slow time course (tau = 2-19 s). This slow time course has precluded the observation of this property in previous studies that used shorter pulses. Contrary to the classical view of FS neurons functional properties, long-duration discharges were followed by a slow afterhyperpolarization lasting < or =23 s. During this postadaptation period, the excitability of the neurons was decreased on average for 16.7 +/- 6.8 s, therefore rendering the cell less responsive to subsequent afferent inputs. Slow adaptation is also reported here for FS neurons recorded in vivo. This longer time scale of adaptation in FS neurons may be critical for balancing excitation and inhibition as well as for the understanding of cortical network computations.

Synaptojanin 1 contributes to maintaining the stability of GABAergic transmission in primary cultures of cortical neurons.

Inhibitory synapses in the CNS can exhibit a considerable stability of neurotransmission over prolonged periods of high-frequency stimulation. Previously, we showed that synaptojanin 1 (SJ1), a presynaptic polyphosphoinositide phosphatase, is required for normal synaptic vesicle recycling (Cremona et al., 1999). We asked whether the stability of inhibitory synaptic responses was dependent on SJ1. Whole-cell patch-clamp recordings of unitary IPSCs were obtained in primary cortical cultures between cell pairs containing a presynaptic, fast-spiking inhibitory neuron (33.5-35 degrees C). Prolonged presynaptic stimulation (1000 stimuli, 2-20 Hz) evoked postsynaptic responses that decreased in size with a bi-exponential time course. A fast component developed within a few stimuli and was quantified with paired-pulse protocols. Paired-pulse depression (PPD) appeared to be independent of previous GABA release at intervals of >/=100 msec. The characteristics of PPD, and synaptic depression induced within the first approximately 80 stimuli in the trains, were unaltered in SJ1-deficient inhibitory synapses. A slow component of depression developed within hundreds of stimuli, and steady-state depression showed a sigmoidal dependence on stimulation frequency, with half-maximal depression at 6.0 +/- 0.5 Hz. Slow depression was increased when release probability was augmented, and there was a small negative correlation between consecutive synaptic amplitudes during steady-state depression, consistent with a presynaptic depletion process. Slow depression was increased in SJ1-deficient synapses, with half-maximal depression at 3.3 +/- 0.9 Hz, and the recovery was retarded approximately 3.6-fold. Our studies establish a link between a distinct kinetic component of physiologically monitored synaptic depression and a molecular modification known to affect synaptic vesicle reformation.

On the cellular and network bases of epileptic seizures.

The highly interconnected networks of the mammalian forebrain can generate a wide variety of synchronized activities, including those underlying epileptic seizures, which often appear as a transformation of otherwise normal brain rhythms. The cerebral cortex and hippocampus are particularly prone to the generation of the large, synchronized bursts of activity underlying many forms of seizures owing to strong recurrent excitatory connections, the presence of intrinsically burst-generating neurons, ephaptic interactions among closely spaced neurons, and synaptic plasticity. The simplest form of epileptiform activity in these structures is the interictal spike, a synchronized burst of action potentials generated by recurrent excitation, followed by a period of hyperpolarization, in a localized pool of pyramidal neurons. Seizures can also be generated in response to a loss of balance between excitatory and inhibitory influences and can take the form of either tonic depolarizations or repetitive, rhythmic burst discharges, either as clonic or spike-wave activity, again mediated both by intrinsic membrane properties and synaptic interactions. The interaction of the cerebral cortex and the thalamus, in conjunction with intrathalamic communication, can also generate spike waves similar to those occurring during human absence seizure discharges. Although epileptic syndromes and their causes are diverse, the cellular mechanisms of seizure generation appear to fall into only two categories: rhythmic or tonic "runaway" excitation or the synchronized and rhythmic interplay between excitatory and inhibitory neurons and membrane conductances.

Brain calculus: neural integration and persistent activity.

Tank and colleagues make in vivo cellular recordings from neurons in a "neural integrator" of the goldfish involved in maintaining eye position. In this circuit, "working" memory may be the result of persistent changes in the state of the local network.

Cellular and network mechanisms of rhythmic recurrent activity in neocortex.

The neocortex generates periods of recurrent activity, such as the slow (0.1-0.5 Hz) oscillation during slow-wave sleep. Here we demonstrate that slices of ferret neocortex maintained in vitro generate this slow (< 1 Hz) rhythm when placed in a bathing medium that mimics the extracellular ionic composition in situ. This slow oscillation seems to be initiated in layer 5 as an excitatory interaction between pyramidal neurons and propagates through the neocortex. Our results demonstrate that the cerebral cortex generates an 'up' or depolarized state through recurrent excitation that is regulated by inhibitory networks, thereby allowing local cortical circuits to enter into temporarily activated and self-maintained excitatory states. The spontaneous generation and failure of this self-excited state may account for the generation of a subset of cortical rhythms during sleep.

Ionic mechanisms underlying repetitive high-frequency burst firing in supragranular cortical neurons.

Neocortical neurons in awake, behaving animals can generate high-frequency (>300 Hz) bursts of action potentials, either in single bursts or in a repetitive manner. Intracellular recordings of layer II/III pyramidal neurons were obtained from adult ferret visual cortical slices maintained in vitro to investigate the ionic mechanisms by which a subgroup of these cells generates repetitive, high-frequency burst discharges, a pattern referred to as "chattering." The generation of each but the first action potential in a burst was dependent on the critical interplay between the afterhyperpolarizations (AHPs) and afterdepolarizations (ADPs) that followed each action potential. The spike-afterdepolarization and the generation of action potential bursts were dependent on Na(+), but not Ca(2+), currents. Neither blocking of the transmembrane flow of Ca(2+) nor the intracellular chelation of free Ca(2+) with BAPTA inhibited the generation of intrinsic bursts. In contrast, decreasing the extracellular Na(+) concentration or pharmacologically blocking Na(+) currents with tetrodotoxin, QX-314, or phenytoin inhibited bursting before inhibiting action potential generation. Additionally, a subset of layer II/III pyramidal neurons could be induced to switch from repetitive single spiking to a burst-firing mode by constant depolarizing current injection, by raising extracellular K(+) concentrations, or by potentiation of the persistent Na(+) current with the Na(+) channel toxin ATX II. These results indicate that cortical neurons may dynamically regulate their pattern of action potential generation through control of Na(+) and K(+) currents. The generation of high-frequency burst discharges may strongly influence the response of postsynaptic neurons and the operation of local cortical networks.

Corticothalamic inputs control the pattern of activity generated in thalamocortical networks.

Absence seizures (3-4 Hz) and sleep spindles (6-14 Hz) occur mostly during slow-wave sleep and have been hypothesized to involve the same corticothalamic network. However, the mechanism by which this network transforms from one form of activity to the other is not well understood. Here we examine this question using ferret lateral geniculate nucleus slices and stimulation of the corticothalamic tract. A feedback circuit, meant to mimic the cortical influence in vivo, was arranged such that thalamic burst firing resulted in stimulation of the corticothalamic tract. Stimuli were either single shocks to mimic normal action potential firing by cortical neurons or high-frequency bursts (six shocks at 200 Hz) to simulate increased cortical firing, such as during seizures. With one corticothalamic stimulus per thalamic burst, 6-10 Hz oscillations resembling spindle waves were generated. However, if the stimulation was a burst, the network immediately transformed into a 3-4 Hz paroxysmal oscillation. This transition was associated with a strong increase in the burst firing of GABAergic perigeniculate neurons. In addition, thalamocortical neurons showed a transition from fast (100-150 msec) IPSPs to slow ( approximately 300 msec) IPSPs. The GABA(B) receptor antagonist CGP 35348 blocked the slow IPSPs and converted the 3-4 Hz paroxysmal oscillations back to 6-10 Hz spindle waves. Conversely, the GABA(A) receptor antagonist picrotoxin blocked spindle frequency oscillations resulting in 3-4 Hz oscillations with either single or burst stimuli. We suggest that differential activation of thalamic GABA(A) and GABA(B) receptors in response to varying corticothalamic input patterns may be critical in setting the oscillation frequency of thalamocortical network interactions.

Membrane mechanisms underlying contrast adaptation in cat area 17 in vivo.

Contrast adaptation is a psychophysical phenomenon, the neuronal bases of which reside largely in the primary visual cortex. The cellular mechanisms of contrast adaptation were investigated in the cat primary visual cortex in vivo through intracellular recording and current injections. Visual cortex cells, and to a much less extent, dorsal lateral geniculate nucleus (dLGN) neurons, exhibited a reduction in firing rate during prolonged presentations of a high-contrast visual stimulus, a process we termed high-contrast adaptation. In a majority of cortical and dLGN cells, the period of adaptation to high contrast was followed by a prolonged (5-80 sec) period of reduced responsiveness to a low-contrast stimulus (postadaptation suppression), an effect that was associated, and positively correlated, with a hyperpolarization of the membrane potential and an increase in apparent membrane conductance. In simple cells, the period of postadaptation suppression was not consistently associated with a decrease in the grating modulated component of the evoked synaptic barrages (the F1 component). The generation of the hyperpolarization appears to be at least partially intrinsic to the recorded cells, because the induction of neuronal activity with the intracellular injection of current resulted in both a hyperpolarization of the membrane potential and a decrease in the spike response to either current injections or visual stimuli. Conversely, high-contrast visual stimulation could suppress the response to low-intensity sinusoidal current injection. We conclude that control of the membrane potential by intrinsic neuronal mechanisms contributes importantly to the adaptation of neuronal responsiveness to varying levels of contrast. This feedback mechanism, internal to cortical neurons, provides them with the ability to continually adjust their responsiveness as a function of their history of synaptic and action potential activity.

Cellular mechanisms of long-lasting adaptation in visual cortical neurons in vitro.

The cellular mechanisms of spike-frequency adaptation during prolonged discharges and of the slow afterhyperpolarization (AHP) that follows, as occur in vivo with contrast adaptation, were investigated with intracellular recordings of cortical neurons in slices of ferret primary visual cortex. Intracellular injection of 2 Hz sinusoidal or constant currents for 20 sec resulted in a slow (tau = 1-10 sec) spike-frequency adaptation, the degree of which varied widely among neurons. Reducing either [Ca(2+)](o) or [Na(+)](o) reduced the rate of spike-frequency adaptation. After the prolonged discharge was a slow (12-75 sec) AHP that was associated with an increase in membrane conductance and a rightward shift in the discharge frequency versus injected current relationship. The reversal potential of the slow AHP was sensitive to changes in [K(+)](o), indicating that it was mediated by a K(+) current. Blockade of transmembrane Ca(2+) conductances did not reduce the slow AHP. In contrast, reductions of [Na(+)](o) reduced the slow AHP, even in the presence of pronounced Ca(2+) spikes. We suggest that the activation of Na(+)-activated and Ca(2+)-activated K(+) currents plays an important role in prolonged spike-frequency adaptation and therefore may contribute to contrast adaptation and other forms of adaptation in the visual system in vivo.

Essential role of phosphoinositide metabolism in synaptic vesicle recycling.

Growing evidence suggests that phosphoinositides play an important role in membrane traffic. A polyphosphoinositide phosphatase, synaptojanin 1, was identified as a major presynaptic protein associated with endocytic coated intermediates. We report here that synaptojanin 1-deficient mice exhibit neurological defects and die shortly after birth. In neurons of mutant animals, PI(4,5)P2 levels are increased, and clathrin-coated vesicles accumulate in the cytomatrix-rich area that surrounds the synaptic vesicle cluster in nerve endings. In cell-free assays, reduced phosphoinositide phosphatase activity correlated with increased association of clathrin coats with liposomes. Intracellular recording in hippocampal slices revealed enhanced synaptic depression during prolonged high-frequency stimulation followed by delayed recovery. These results provide genetic evidence for a crucial role of phosphoinositide metabolism in synaptic vesicle recycling.

Modulation of a pacemaker current through Ca(2+)-induced stimulation of cAMP production.

Brief increases in [Ca2+]i can result in prolonged changes in neuronal properties. A Ca(2+)-dependent modulation of the hyperpolarization-activated cation current (Ih) controls the slow recurrence of synchronized thalamocortical activity. Here we show that the persistent activation of Ih is initiated by rapidly increased [Ca2+]i and subsequent production of cAMP. The modulation is maintained via a facilitated interaction of cAMP with open (voltage-gated) h-channels, inducing prolonged activation of Ih that may outlast the presence of increased free [Ca2+]i and [cAMP]i. This persistent Ih activation may control the presence and periodicity of both normal and abnormal synchronized thalamocortical rhythms.

Dynamic properties of corticothalamic excitatory postsynaptic potentials and thalamic reticular inhibitory postsynaptic potentials in thalamocortical neurons of the guinea-pig dorsal lateral geniculate nucleus.

The properties of postsynaptic potentials evoked by stimulation of cortical, retinal and GABAergic thalamic afferents were examined in vitro in thalamocortical neurons of the guinea-pig dorsal lateral geniculate nucleus. Brief trains of stimulation (2-10 stimuli) delivered to corticothalamic fibers led to a frequency-dependent increase in excitatory postsynaptic potential amplitude associated with an increase in activation of both N-methyl-D-aspartate and non-N-methyl-D-aspartate glutamate receptors. In addition, repetitive stimulation of corticothalamic fibers also gave rise to a slow excitatory postsynaptic potential that was blocked by local application of the glutamate metabotropic receptor antagonist alpha-methyl-4-carboxyphenylglycine. In contrast, repetitive stimulation of optic tract fibers resulted in monosynaptic excitatory postsynaptic potentials that did not potentiate and were not followed by the generation of a slow excitatory postsynaptic potential. Repetitive activation of the optic radiation also evoked both GABA(A) and GABA(B) receptor-mediated inhibitory postsynaptic potentials. These inhibitory postsynaptic potentials exhibited frequency-dependent depression during repetitive activation. The presence of frequency-dependent facilitation of corticothalamic excitatory postsynaptic potentials and frequency-dependent decrement of inhibitory postsynaptic potentials, as well as the ability of corticothalamic fibers to activate glutamate metabotropic receptors, suggests that sustained activation of corticothalamic afferents in vivo may result in postsynaptic responses in thalamocortical cells that are initially dominated by GABAergic inhibitory postsynaptic potentials followed by prominent monosynaptic excitatory postsynaptic potentials as well as a slow depolarization of the membrane potential.Therefore, the corticothalamic system may inhibit or enhance the excitability and responsiveness of thalamocortical neurons, based both on the spatial and temporal features of thalamocortical interactions.

The functional influence of burst and tonic firing mode on synaptic interactions in the thalamus.

Thalamocortical and perigeniculate (PGN) neurons can generate action potentials either as Ca2+ spike-mediated high-frequency bursts or as tonic trains. Using dual intracellular recordings in vitro in monosynaptically connected pairs of PGN and dorsal lateral geniculate nucleus (LGNd) neurons, we found that the functional effect of synaptic transmission between these cell types was strongly influenced by the membrane potential and hence the firing mode of both the pre- and postsynaptic neurons. Activation of single action potentials or low-frequency spike trains in PGN or thalamocortical neurons resulted in the generation of PSPs that were 0.5-2.0 mV in amplitude. In contrast, the generation of Ca2+ spike-mediated bursts of action potentials in the presynaptic cell increased these PSPs to an average of 4.4 mV for the IPSP and 3.0 mV for the EPSP barrage, because of temporal summation and/or facilitation. If the postsynaptic neuron was at a resting membrane potential (e.g., -65 mV), these PSP barrages could result in the activation of a low-threshold Ca2+ spike and burst of action potentials. These results demonstrate that the burst firing mode of action potential generation is a particularly effective means by which perigeniculate and thalamocortical neurons may influence one another. We propose that the activation of burst discharges in these cell types is essential for the generation of some forms of synchronized rhythmic oscillations of sleep and of epileptic seizures.

Functional and ionic properties of a slow afterhyperpolarization in ferret perigeniculate neurons in vitro.

Intracellular recordings from spontaneously spindling GABAergic neurons of the ferret perigeniculate nucleus in vitro revealed a fast afterhyperpolarization after each action potential, a medium-duration afterhyperpolarization after each low-threshold Ca2+ spike, and a slow afterhyperpolarization after the cessation of spindle waves. The slow afterhyperpolarization was associated with an increase in membrane conductance, and the reversal potential was sensitive to extracellular [K+]o, indicating that it is mediated at least in part by the activation of a K+ conductance. However, the block of Ca2+ channels did not block the slow afterhyperpolarization, whereas the block of Na+ channels did block this event, even after the generation of repetitive Ca2+ spikes, indicating that it is mediated by a Na+-activated K+ current. Application of apamin reduced the afterhyperpolarization and enhanced a plateau potential after each low-threshold Ca2+ spike. This plateau potential could result in a prolonged depolarization of perigeniculate neurons, even before the application of apamin, resulting in the generation of tonic discharge. The plateau potential was blocked by the local application of tetrodotoxin, indicating that it is mediated by a persistent Na+ current. The activation and interaction of these slowly developing and persistent currents contributes significantly to low-frequency components of spindle wave generation. In particular, we suggest that the activation of the slow afterhyperpolarization may contribute to the generation of the spindle wave refractory period in vitro.

Periodicity of thalamic spindle waves is abolished by ZD7288,a blocker of Ih.

The actions of the novel bradycardiac agent ZD7288 [4-(N-ethyl-N-phenylamino)-1, 2-dimethyl-6-(methylamino)pyrimidinium chloride] were investigated on the hyperpolarization-activated cation current Ih and on network activity in spontaneously spindling ferret lateral geniculate (LGNd) slices in vitro using intracellular recording techniques. In voltage-clamp recordings, local application of ZD7288 (1 mM in micropipette) resulted in a complete block of Ih, whereas in current-clamp recordings, application of this agent resulted in an abolition of the depolarizing sag activated by hyperpolarization and decreased the frequency of intrinsic delta-oscillations for which Ih acts as a pacemaker current. In addition, block of Ih with ZD7288 resulted in an abolition of the afterdepolarization (ADP) that follows repetitive hyperpolarization and rebound burst firing as well as that occurring in between spindle waves. The block of the ADP was associated with a block of the spindle wave refractory period such that continuous 6- to 10-Hz oscillations were generated throughout the network. These findings give further support to the hypothesis that Ih is critically involved in the generation of slow rhythmicity in synchronized thalamic activity.