Extrusion of Intracellular Calcium Ion After In Vitro Ischemia in the Rat Hippocampal CA1 Region

Simultaneous recordings of intracellular Ca2+([Ca2+]i) signal and extracellular DC potential were obtained from the CA1 region in 1-[6-amino-2-(5-carboxy-2-oxazolyl)-5-benzofuranyloxy]-2-(2-amino-5-methylphenoxy)-ethane- N, N, N′, N′-tetraacetic acid penta-acetoxymethyl ester (Fura-2/AM)-loaded rat hippocampal slices. Superfusion with oxygen- and glucose-deprived medium (in vitro ischemia) for 5–6 min produced a rapid rise of the [Ca2+]i level in the stratum radiatum (rising phase of the [Ca2+]i signal), which occurred simultaneously with a rapid negative DC potential (rapid negative potential). When oxygen and glucose were reintroduced, the increased [Ca2+]i signal diminished rapidly (falling phase of the [Ca2+]i signal) during the generation of a slow negative DC potential (slow negative potential), which occurred within 1 min from the onset of the reintroduction. Thereafter, the [Ca2+]i signal partially and the slow negative potential completely returned to the preexposure level approximately 6 min after the reintroduction. The changes in [Ca2+]i signal during and after in vitro ischemia were very similar to the changes in the membrane potential of glial cells. The rising and falling phases of [Ca2+]i signal corresponded to the rapid depolarization and a depolarizing hump, respectively, in the repolarizing phase of glial cells. A prolonged application of in vitro ischemia or a reintroduction of either glucose or oxygen suppressed the falling phase after ischemic exposure. The application of ouabain (30 μM) generated both a rapid negative potential and a rapid elevation of [Ca2+]i, but no slow negative potential or rapid reduction in [Ca2+]i were observed. When oxygen and glucose were reintroduced to slices in the Na+-free or ouabain- or Ni2+-containing medium, the falling phase was suppressed. The falling phase was significantly accelerated in Ca2+- and Mg2+-free with EGTA-containing medium. In contrast, the falling phase was significantly slower in the Ca2+-free with high Mg2+- and EGTA-containing medium. The falling phase of the [Ca2+]isignal after ischemic exposure is thus considered to be primarily dependent on the reactivation of Na+, K+-ATPases, while the extrusion of cytosolic Ca2+ via the forward-mode operation of Na+/Ca2+ exchangers in glial cells is thought to be directly involved in the rapid reduction of [Ca2+]i after ischemic exposure.

Spontaneous ictal-like discharges and sustained potential shifts in the developing rat neocortex

1. Intra- and extracellular recording techniques were used to study epileptogenesis in in vitro slices of immature rat neocortex. Slices of sensorimotor cortex were prepared from animals 5-60 days old. Epileptiform activity was induced by bath application of 50 microM picrotoxin. 2. Convulsant-induced paroxysmal activity was observed only rarely in the youngest age group (5-7 days) and consisted of orthodromically evoked bursts of low-amplitude isolated discharges. This activity was labile and could be evoked only at long interstimulus intervals (greater than 10 s). 3. Extracellular recordings in slices from 8- to 15-day-old rats showed spontaneous epileptiform activity consisting of 10- to 30-s paroxysms of repetitive spike discharges superimposed on a 3- to 5-mV negative steady potential. This steady potential declined slightly during the course of the prolonged discharge and returned quickly to base line following the last spike discharge. 4. Laminar analysis of epileptiform activity in 8- to 15-day-old rats showed that the spike discharges were negative and superimposed on a positive slow wave in superficial cortical layers. At 100 micron below the pial surface, the slow potential reversed polarity and remained negative throughout the remainder of the cortex. Spike discharges reversed polarity 800 micron below the pial surface. 5. In intracellular recordings from slices obtained from 9- to 14-day-old animals, each paroxysm began with a sharply rising membrane depolarization (paroxysmal depolarizing shift, or PDS). A second PDS occurred before the cells repolarized to their resting potential. A series of PDSs then followed, superimposed on a sustained membrane depolarization. This was associated with a 33% decrease in input resistance. Afterhyperpolarizations (AHPs) following termination of the depolarization were low in amplitude or absent. 6. In the 16- to 30-day-old age group, extracellular recordings showed paroxysmal activity consisting of 3-10 initial spikes followed by a sustained, slow, negative-potential shift. This slow potential could be as great as 30 mV in amplitude and could last as long as 180 s. Paroxysmal events recurred spontaneously at intervals of 4-11 min. Spontaneous PDSs and slow, negative-potential shifts were not observed after 30 days of age, although PDSs could still be evoked by orthodromic stimulation. 7. Intracellular recordings in the 16- to 30-day-old group revealed that each paroxysmal event consisted of an initial period of increased synaptic activity and cellular firing, followed by a marked, long-lasting depolarization (LLD), culminating in an AHP.(ABSTRACT TRUNCATED AT 400 WORDS)