Many patients develop temporal lobe epilepsy after trauma, but basic mechanisms underlying the development of chronic seizures after head injury remain poorly understood. Using the controlled cortical impact injury model we examined whether mice developed spontaneous seizures after mild (0.5mm injury depth) or severe (1.0mm injury depth) brain injury and how subsequent posttraumatic mossy fiber sprouting was associated with excitability in the dentate gyrus 42–71 days after injury. After several weeks, spontaneous behavioral seizures were observed in 20% of mice with mild and 36% of mice with severe injury. Mossy fiber sprouting was typically present in septal slices of the dentate gyrus ipsilateral to the injury, but not in control mice. In slices with mossy fiber sprouting, perforant path stimulation revealed a significant reduction (P<0.01) in paired-pulse ratios in dentate granule cells at 20ms and 40ms interpulse intervals, but not at 80ms or 160ms intervals. These slices were also characterized by spontaneous and hilar-evoked epileptiform activity in the dentate gyrus in the presence of Mg2+-free ACSF containing 100μM picrotoxin. In contrast, paired-pulse and hilar-evoked responses in slices from injured animals that did not display mossy fiber sprouting were not different from controls. These data suggest the development of spontaneous posttraumatic seizures as well as structural and functional network changes associated with temporal lobe epilepsy in the mouse dentate gyrus by 71d after CCI injury. Identifying experimental injury models that exhibit similar pathology to injury-induced epilepsy in humans should help to elucidate the mechanisms by which the injured brain becomes epileptic.

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PRV-Bartha ͉ PRV 152 ͉ suprachiasmatic nucleus ͉ retinal ganglion cell T he use of neurotropic alphaherpesviruses has greatly advanced our ability to visualize ensembles of neurons that contribute to multisynaptic circuits in the central nervous system (CNS) (1). In particular, the attenuated vaccine strain of pseudorabies virus (PRV-Bartha) has been used successfully as a self-amplifying neural tracer after peripheral application or direct injection into brain parenchyma (2-4). The usefulness of PRV as a neural tracer relies on its ability to infect chains of hierarchically connected neurons via specific transsynaptic passage of progeny virus rather than infection by lytic release into the extracellular space (4, 5). Typically, PRV infects the CNS by invading neurons in the periphery and then replicating and spreading to the CNS via synaptically linked neurons. However, PRV can also invade neurons through their somata if the viral concentration is sufficient (6), as evidenced by primary infection of retinal ganglion cells (RGCs) after intravitreal injection of PRV (7-9). Infection of RGCs with PRV-Bartha, followed by viral replication, results in the anterograde transsynaptic infection of a restricted set of retinorecipient neurons [i.e., suprachiasmatic nucleus (SCN), intergeniculate leaflet (IGL), pretectum (PT), and lateral terminal nucleus]. Intravitreal injection of the wild-type virus, PRV-Becker, produces transneuronal infection of neurons in all retinorecipient subcortical regions (7). The factors that determine the specificity of PRV-Bartha infection of selective retinorecipient targets are not completely understood, although deletion of specific genes in PRV-Becker results in a restricted neurotropism identical to that demonstrated with PRV-Bartha (10).Although viral transsynaptic tracing represents an important methodological advance for the analysis of CNS circuits, functional analysis of virus-infected neurons has been limited to sensory or sympathetic ganglia in culture (11, 12) because of the inability to identify virus-infected neurons in situ. Analyses of electrophysiological properties of neurons, in the context of known functional connections of the recorded neuron, would represent a further important methodological advance for the analysis of CNS circuits.The development of retrogradely transported fluorescent tracers has allowed investigators to examine the physiology of neurons with known projections (13-15). However, such studies usually require direct and accurate injection of a target region followed by retrograde transport to identify first-order neurons projecting to the target. Other ''prelabeling'' studies have used constructs of green fluorescent protein to label neurons that possess a particular genetic phenotype, such as expression of gonadotropin-releasing hormone (16). Both of these techniques have allowed examination of the physiological properties of neurons in vitro that possess presumed anatomical or functional correlates in the intact animal.We now report a neuron-labeling...

1. Intracellular recordings were made from supraoptic neurones in vitro from hypothalamic explants prepared from adult male rats. Neurones were injected with biotinylated markers, and of thirty-nine labelled neurones, nineteen were identified immunocytochemically as containing oxytocin-neurophysin and twenty as containing vasopressin-neurophysin. 2. Vasopressin and oxytocin neurones did not differ in their resting membrane potential, input resistance, membrane time constant, action potential height from threshold, action potential width at half-amplitude, and spike hyperpolarizing after-potential amplitude. Both cell types exhibited spike broadening during brief, evoked spike trains (6-8 spikes), but the degree of broadening was slightly greater for vasopressin neurones. When hyperpolarized below -75 mV, all but one neurone exhibited a transient outward rectification to depolarizing pulses, which delayed the occurrence of the first spike.3. Both cell types exhibited a long after-hyperpolarizing potential (AHP) following brief spike trains evoked either with a square wave pulse or using 5 ms pulses in a train. There were no significant differences between cell types in the size of the AHP evoked with nine spikes, or in the time constant of its decay. The maximal AHP evoked by a 180 ms pulse was elicited by an average of twelve to thirteen spikes, and neither the size of this maximal AHP nor its time constant of decay were different for the two cell types.4. In most oxytocin and vasopressin neurones the AHP, and concomitantly spike frequency adaptation, were markedly reduced by the bee venom apamin and by d-tubocurarine, known blockers of a Ca2+-mediated K+ conductance. However, a minority of neurones, of both cell types, were relatively resistant to both agents.