Repetitive Spreading Depression-Like Events Result in Cell Damage in Juvenile Hippocampal Slice Cultures Maintained in Normoxia

Abstract

Prolonged seizures, e.g., induced by fever, experienced early in life are considered a precipitating injury for the subsequent development of temporal lobe epilepsy. During in vitro epileptiform activity, spreading depressions (SDs) have often been observed. However, their contribution to changes in the properties of juvenile neuronal tissue is unknown. We therefore used the juvenile hippocampal slice culture preparation (JHSC) maintained in normoxia (20% O(2)-5% CO(2)-75% N(2)) to assess the effect of repetitive SD-like events (SDLEs) on fast field potentials and cell damage. Repetitive SDLEs in the CA1 region could be induced in about two-thirds of the investigated JHSCs (n = 61) by repetitive electrical stimulation with 2-200 pulses. SDLEs were characterized by a transient large negative field potential shift accompanied by intracellular depolarization, ionic redistribution, slow propagation (assessed by intrinsic optical signals) and glutamate receptor antagonist sensitivity. The term "SDLE" was used because evoked fast field potentials were only incompletely suppressed and superimposed discharges occurred. With 20 +/- 1 repetitive SDLEs (interval of 10-15 min, n = 7 JHSCs), the events got longer, their amplitude of the first peak declined, while threshold for induction became reduced. Evoked fast field potentials deteriorated and cell damage (assessed by propidium iodide fluorescence) occurred, predominantly in regions CA1 and CA3. As revealed by measurements of tissue partial oxygen pressure during SDLEs repetitive transient anoxia accompanying SDLE might be critical for the observed cell damage. These results, limited so far to the slice culture preparation, suggest SDs to be harmful events in juvenile neuronal tissue in contrast to what is known about their effect on adult neuronal tissue.

Differential Effects of Hypothermia on Early and Late Epileptiform Events After Severe Hypoxia in Preterm Fetal Sheep

“…Intriguingly, there is in vitro evidence that the juvenile brain is more likely than the adult to show spontaneous spreading depression-like events during recovery from hypoxia (Luhmann and Kral 1997). Further, whereas these events are not injurious to the normoxic adult brain (Somjen 2001), in juvenile normoxic hippocampal slice cultures, repetitive spreading depression-like events led to deterioration of evoked fast field potentials and cell damage (Pomper et al 2006).…”

Gamma Oscillations and Spontaneous Network Activity in the Hippocampus Are Highly Sensitive to Decreases in pO2and Concomitant Changes in Mitochondrial Redox State

“…Fluorescence and pO 2 recordings were made in submerged recording chambers mounted on Axioskop (Zeiss, Jena, Germany) and BX51WI upright microscopes (Olympus, Hamburg, Germany) using 20ϫ [0.5 numerical aperture (NA)] and 10ϫ (0.3 NA) water-immersion objectives or a 4ϫ objective (0.28 NA) with a water cap. For recordings of multiunit activity and gamma oscillations, self-made interface recording chambers were used (Stenkamp et al, 2001;Pomper et al, 2006). Recording microelectrodes (K ϩ -sensitive/local field potential, O 2 -sensitive) were placed in stratum pyramidale of the CA3 subfield, and a bipolar stimulation electrode (tungsten filament, tip diameter of ϳ10 m) was positioned close to the dentate gyrus to activate fiber tracts to CA3.…”

“…An oxygen sensor microelectrode (tip diameter of 10 m; 737GC; Diamond General Development, Ann Arbor, MI) was used to continuously measure changes in pO 2 . This new type of modified polarographic Clark electrodes consists of a glass-insulated Ag/ AgCl reference anode and a guard cathode with the advantages of low sensitivity to motion artifact, a time constant of Ͻ1 s (0 -90%), minimal interaction with tissue, and low O 2 consumption (Foster et al, 2005;Pomper et al, 2006;Takano et al, 2007). For polarization, the electrode was connected to a picoammeter and the tip was maintained in nongassed ACSF overnight, which resulted in a stable current readout.…”

“…The Clark-style microelectrode with low O 2 consumption has been commonly used to monitor interstitial pO 2 levels even under hypoxic conditions in hippocampal slice preparations (Foster et al, 2005;Pomper et al, 2006) and in vivo (Takano et al, 2007). We determined a pO 2 of 578 Ϯ 17 mmHg (n ϭ 9) at the surface of the slices (Fig.…”

Reentrant spiral waves of spreading depression cause macular degeneration in hypoglycemic chicken retina

“…The finding that during SD the tissue oxygen level is comparable to the in vivo situation (31,32) indicates that hypoxia is not a cause of SD initiation or subsequent macular degeneration. Further, lack of transient anoxia during SDs does not support the suggestion that occurrence of anoxic periods may be a compromising factor leading to neuronal damage (33). Spontaneous SD activity is increased by hyposmolarity, which is ascribable to reduced extracellular space and greater accumulation of K + and possibly neurotransmitters.…”