Spread of epileptiform potentials in the neocortical slice: recordings with voltage-sensitive dyes.

The spread of epileptiform potentials in guinea pig neocortical slices was investigated by use of voltage sensitive dyes and a fast optical recording technique. Epileptiform activity was induced in a perfusion medium containing 10-20 microM bicuculline-methiodide and by single pulse stimulation of layer I or the white matter. The location of minimal and maximal amplitudes, the shape of the potentials at specific sites and the velocity of spread were independent from the specific stimulation site. The expression of epileptiform activity appeared to depend on specific, possibly geometrical, properties of the tissue.

Partial suppression of GABAA-mediated inhibition induces spatially restricted epileptiform activity in guinea pig neocortical slices.

The spatio-temporal distribution of evoked activity in guinea pig neocortical slices was investigated during partial suppression of gamma-aminobutyric acid (GABA)A-mediated synaptic inhibition with different concentrations of bicuculline. Activity patterns were recorded by use of a voltage-sensitive dye and a fast optical recording technique. At non-epileptogenic concentrations of bicuculline (0.6-2.5 microM), evoked potentials were of longer duration and larger amplitude, but the spatial extent of spread in the horizontal direction was unaffected. At threshold epileptogenic concentrations of bicuculline (1.25-5 microM), spatially restricted epileptiform activity developed at a distance from the stimulation site which was clearly separated from potentials with non-epileptic characteristics close to the stimulation site. It is concluded that, under moderate disinhibition, stimulus-evoked activity has a suppressive effect on spread and development of epileptiform activity, probably through synchronous activation of still-functioning inhibitory circuits.

Spatio-temporal distribution of epileptiform activity in slices from human neocortex: recordings with voltage-sensitive dyes.

The spatio-temporal distribution of epileptiform activity was investigated in slices from human temporal neocortex resected during epilepsy surgery. Activity was recorded by use of a voltage-sensitive dye and an optical recording system. Epileptiform activity was induced with 10 microM bicuculline and electrical stimulation of layer I. In 10 slices from six patients investigated, epileptiform activity spread across most of the slice. Largest amplitudes were located in layer II/III. Epileptiform activity was characterized by long-lasting potentials with slow rising phases and a low velocity of spread in the horizontal direction (0.044 m/s). This spatio-temporal pattern of epileptiform activity in human slices was similar to that found previously in neocortical slices from guinea pigs with bicuculline. In four of nine human slices investigated under control bath conditions (in non-epileptogenic medium), the spatio-temporal activity patterns were similar to those of guinea pigs in non-epileptogenic medium. In the remaining five human slices, however, the spread in the horizontal direction was significantly larger (4188 microm) in non-epileptogenic medium than that found in slices from guinea pigs (2171 microm). Activity in human slices showing such 'wide spread' in control bath conditions occasionally had characteristic features of epileptiform activity. Further work will have to clarify whether these epileptiform features reflect intrinsic epileptiform properties in human tissue slices.

The contribution of intracortical connections to horizontal spread of activity in the neocortex as revealed by voltage sensitive dyes and a fast optical recording method.

Coronal slices from guinea-pig visual neocortex were stained with voltage-sensitive fluorescence dyes RH414 or RH795. Activity was evoked by electrical stimulation of either the white matter or layer I. Emitted light intensity changes representing summated changes of membrane potential were recorded by a 10 x 10 photodiode array with a temporal resolution of 0.4 ms and a spatial resolution of 94 microns. The distribution and spread of activity in the horizontal direction was analysed. Following stimulation of the white matter or layer I, two regions of activity were differentiated in the medio-lateral direction: a central region (approximately 1 mm wide) of high-amplitude activity close to the stimulation electrode and, distant from the stimulation electrode, peripheral regions of low-amplitude activity. Central and peripheral regions differed in their rates of decline, their relative extent with stimulation of different sites and within different layers. The total extent of non-synaptic evoked activity did not exceed that of the central region of high-amplitude activity. Along the extent of non-synaptic activity, onset latencies of potentials were almost constant. Thus, activity of high amplitude in the central region was likely mediated by simultaneous activation of distributed afferent fibres. In contrast, no non-synaptic activity was found in peripheral regions. Therefore it is suggested that this low-amplitude activity was mediated without direct afferent activation but via long-distance intracortical horizontal pathways. These pathways are known to terminate in layer III, and accordingly latencies of responses in the periphery were shortest in upper cortical layers, whereas in the central region, latencies increased from lower to upper cortical layers.

Epileptiform activity in the guinea-pig neocortical slice spreads preferentially along supragranular layers--recordings with voltage-sensitive dyes.

The spread of epileptiform activity was monitored in guinea-pig neocortical slices by the use of a voltage-sensitive dye (RH795) and a fast optical recording technique. Epileptiform activity induced by bicuculline methiodide (10-20 microM) and single-pulse stimulation spread from the stimulation site in layer I or in the white matter across most of the slice. Different lesions were made in the slice in order to specify the neuronal connections used for spread in the horizontal direction. In the slice, intracortical connections are necessary for the spread of epileptiform activity, as shown by vertical cuts through all cortical layers but sparing the white matter. Horizontal connections were interrupted by cuts parallel to the axis of pyramidal neurons through either supragranular or infragranular layers. Vertical connections were interrupted by cuts perpendicular to the axis of pyramidal neurons separating supragranular and infragranular layers. Spread of epileptiform activity in the horizontal direction was not hindered by horizontal cuts. Vertical cuts through infragranular layers also did not hinder the spread of epileptiform activity. In contrast, vertical cuts through supragranular layers either abolished completely (nine slices) or delayed significantly (ten slices) the spread of epileptiform activity. The mean delay at the supragranular lesion was 44 ms in layer III and 30 ms in layer V; at the infragranular lesion the mean delay was 2 ms in layer III and 6 ms in layer V. Also, with horizontal cuts, in three out of five slices the velocity of spread was significantly lower in infragranular as compared to supragranular layers. It is concluded that both supra- and infragranular layers if isolated possess the ability to initiate and propagate epileptiform activity independently. However, in the intact slice the influence of the supragranular networks on initiation and propagation of epileptiform activity appears to dominate.

Evoked changes of membrane potential in guinea pig sensory neocortical slices: an analysis with voltage-sensitive dyes and a fast optical recording method.

Coronal slices from guinea pig visual neocortex were stained with voltage-sensitive fluorescent dyes RH414 or RH795. Activity was evoked by electrical stimulation of either white matter or layer I. Emitted-light intensity changes representing summated changes of membrane potential were recorded by a 10 x 10 photodiode array with a temporal resolution of 0.4 ms and a spatial resolution of 60 microns or 94 microns. Following either stimulation of layer I or of white matter, maximal activity was located close to the respective stimulation electrode, in upper layer III/II, and between layer IV and V. With stimulation of the white matter, additional peak activity was recorded from upper layer VI. Non-synaptic activity was separated from mixed (synaptic and non-synaptic) activity by comparing responses obtained in standard perfusion medium with those obtained in perfusion medium from which the calcium was omitted, such that synaptic transmission was blocked. With stimulation of the white matter, most of the evoked activity in lower cortical layers was of non-synaptic origin. This non-synaptic activity consisted of early and fast potentials, which were predominant in layer VI and probably represented presynaptic fibre activity, and of slower components that were presumably of antidromic origin. Significant postsynaptic activity was only found in upper layer III/II. In contrast, with stimulation of layer I, most of the evoked activity was of postsynaptic origin. Early and fast non-synaptic potentials consisting of presynaptic fibre activity were confined to layer I. Slower non-synaptic activity, that might reflect direct dendritic activation, was minimal and was confined to upper cortical layers. Thus, following either stimulation of layer I or of white matter, the major postsynaptic components were found in upper layer III/II. It is suggested that the postsynaptic response following stimulation of white matter resulted from di- or polysynaptic activation by afferent fibres. The postsynaptic response to stimulation of layer I was presumably a monosynaptic activation of apical dendrites from pyramidal cells by layer I horizontal fibres. Activity following stimulation of white matter spread faster than activity following stimulation of layer I. This might reflect the difference in active conduction along afferent and efferent fibres on the one hand and in passive conductance along the dendritic tree on the other hand.

Optical recording of epileptiform voltage changes in the neocortical slice.

Voltage sensitive probes were used to monitor the development, distribution, and spread of epileptiform potentials with a photodiode array in neocortical slices of guinea pigs. Epileptiform activity was induced by bath application of bicuculline-methiodide or 3,4-diaminopyridine and electrical stimulation of white matter or cortical layer I. Stimulation evoked a primary or early potential which was followed by a delayed epileptiform potential with a larger spatial extent. Shape, duration and amplitude of the delayed epileptiform potential varied strongly among slices and across the recording area and could reach largest amplitudes at a distance from the stimulation point. At a specific recording site, however, with repeated stimulation, potentials were generated in a stereotyped way. Intracellularly recorded delayed epileptiform potentials corresponded very closely at least to the early part of the optical response. Epileptiform activity appeared in layer III as soon as the primary potential reached sufficient amplitude there. Apart from this relationship, the distribution and spread of maximal amplitudes of delayed epileptiform potentials were segregated from those of early potentials. Early potentials reached maximal amplitudes close to the stimulation site. In contrast, the largest amplitudes of delayed epileptiform potentials were always found in layer III. A second maximum occasionally occurred in layer V. The horizontal amplitude distribution of epileptiform potentials was asymmetric, i.e. amplitudes increased to one side and decreased to the other. Early potential maxima spread from deeper to upper layers when initiated by white matter stimulation and from upper to deeper layers when initiated by layer I stimulation. In contrast, delayed epileptiform potentials always spread from layer III to lower layers and to the sides. Velocity of spread of early potentials and delayed epileptiform potentials differed systematically along the vertical and horizontal axis. The distribution of maximal amplitudes, shape, and pattern, of spread of epileptiform potentials was the same whether white matter or layer I was stimulated. The independence of delayed epileptiform potential characteristics from the point of stimulation and from early potential characteristics suggests that epileptiform activity is determined by intrinsic properties of the cortex and not by afferent activation.

Interictal afterdischarge in focal penicillin epilepsy: thalamocortical unit activity.

The involvement of thalamic versus cortical structures for the initiation and maintenance of brief interictal afterdischarge was evaluated by recording extracellularly units and field potentials from different subcortical and cortical sites. Afterdischarge oscillations at 16 to 22/s that followed interictal spikes with a delay of 170 to 220 ms usually appeared 10 to 30 min after the topical application of penicillin to the cat's precruciate cortex. Units in the ventrolateral nucleus of the thalamus fired in burst discharges during cortical afterdischarge and less reliably during the cortical interictal spike. In contrast, units recorded at the cortical penicillin focus and homologous contralateral site remained silent during afterdischarge but had a typical burst discharge during the interictal spike. Although these data support a thalamic basis for the rhythm, the lack of an afterdischarge-like oscillation in the thalamic field potential and the independent appearance of afterdischarge and cortical recruiting waves elicited by stimulation of the nucleus reticularis would favor its cortical origin. In accordance with its frequency characteristics and data gained from earlier cooling studies we suggest a cortical mechanism requiring thalamic triggering for the generation of afterdischarge.

Spatiotemporal distribution of intracellular calcium transients during epileptiform activity in guinea pig hippocampal slices.

Calcium ions are known to play an important role in epileptogenesis. Although there is clear evidence for increased neuronal calcium influx during epileptiform potentials, direct measurements of the corresponding intracellular calcium transients are rare and the origin of calcium influx is not known. Therefore the spatial and temporal distribution of intracellular calcium transients during epileptiform activity in guinea pig hippocampal slices was monitored with the use of the indicator Calcium-Green and a fast optical recording method. Two models of epilepsy (bicuculline and low Mg2+) were compared. In both models, single epileptiform events were evoked by electrical stimulation of the Schaffer collaterals in CA1 or of stratum pyramidale in area CA3. Intracellular calcium transients during epileptiform activity were approximately 5 times larger than during control stimulation. Calcium transients during epileptiform activity were present across at least the entire CA1 area, whereas presynaptic calcium transients from stimulated fibers were only seen at a distance up to 1 mm from the stimulation site. DL-2-amino-5-phosphonovaleric acid (APV), a specific antagonist of the N-methyl-D-aspartate (NMDA) receptor, abolished low-Mg2+ epileptiform activity and reduced bicuculline-induced epileptiform activity; it reduced calcium transients following stimulation of CA1 by only 29% (bicuculline) and 38% (low Mg2+). For comparison, calcium transients during control stimulation were 78% (bicuculline) and 69% (low Mg2+) smaller than epileptiform calcium transients. At a distance from the stimulation site, calcium transients and their NMDA-receptor-dependent components were largest in stratum pyramidale in the bicuculline model and in stratum oriens in the low-Mg2+ model. In both models, minimal onset latencies of calcium influx shifted with increasing distance to the stimulation electrode from stratum radiatum to stratum oriens. APV reduced the extent of spread of calcium transients in the low-Mg2+ model. In the bicuculline model, the spatial extent of spread of epileptiform calcium transients was not affected by application of APV; however, the mean velocity of spread was reduced from 0.20 to 0.12 m/s. In conclusion, the large size of calcium transients and of their NMDA-receptor-dependent components in stratum pyramidale or stratum oriens as well as shortest onset latencies of calcium transients at these sites suggest an important role of cell somata, basal dendrites, and possibly local circuit excitatory interactions for the generation and spread of epileptiform activity.

Interictal afterdischarge in focal penicillin epilepsy: block by thalamic cooling.

Interictal spikes recorded from a penicillin focus in the precruciate cortex of urethane-anesthetized cats were followed by brief afterdischarge oscillations that occurred at a delay of 170 to 220 ms from the interictal spike and consisted of as many as five cycles at 16 to 22/s. The origin of this afterdischarge was investigated by cooling different subcortical sites and thus blocking them reversibly. At none of the sites did cooling result in a block of the interictal spike whereas cooling of the ventrolateral nucleus of the thalamus resulted in block of the afterdischarge. Other subcortical sites had either no or less reliable effects on it. We conclude that afterdischarge but not the interictal spike depends on thalamic input, either as a generator of the rhythm or as a trigger for a cortically maintained oscillation.

Connections of the anterior ectosylvian visual area (AEV).

We have previously described a visual area situated in the cortex surrounding the deep infolding of the anterior ectosylvian sulcus of the cat (Mucke et al. 1982). Using orthograde and retrograde transport methods we now report anatomical evidence that this anterior ectosylvian visual area (AEV) is connected with a substantial number of both cortical and subcortical regions. The connections between AEV and other cortical areas are reciprocal and, at least in part, topographically organized: the rostral AEV is connected with the bottom region of the presylvian sulcus, the lower bank of the cruciate sulcus, the rostral part of the ventral bank of the splenial sulcus, the rostral portion of the lateral suprasylvian visual area (LS) and the lateral bank of the posterior rhinal sulcus; the caudal AEV is connected with the bottom region of the presylvian sulcus, the caudal part of LS, the ventral part of area 20 and the lateral bank of the posterior rhinal sulcus. Subcortically, AEV has reciprocal connections with the ventral medial thalamic nucleus (VM), with the medial part of the lateralis posterior nucleus (LPm), as well as with the lateralis medialis-suprageniculate nuclear (LM-Sg) complex. These connections are also topographically organized with more rostral parts of AEV being related to more ventral portions of the LPm and LM-Sg complex. AEV also projects to the caudate nucleus, the putamen, the lateral amygdaloid nucleus, the superior colliculus, and the pontine nuclei. It is concluded that AEV is a visual association area which functionally relates the visual with both the motor and the limbic system and that it might play a role in the animal's orienting and alerting behavior.