The mode of death of epilepsy-induced “dark” neurons is neither necrosis nor apoptosis: An electron-microscopic study

Abstract

Morphological aspects of the formation and fate of neurons that underwent dramatic ultrastructural compaction (“dark” neurons) induced by 4-aminopyridine epilepsy were compared in an excitotoxic and a neighboring normal-looking area of the rat brain cortex. In the excitotoxic area, the later the ultrastructural compaction began after the outset of epilepsy, the higher the degree of mitochondrial swelling and ribosomal sequestration were; a low proportion of the affected neurons recovered in 1 day; the others were removed from the tissue through a necrotic-like sequence of ultrastructural changes (swelling of the cell, gradual disintegration of the intracellular organelles and dispersion of their remnants into the surroundings through large gaps in the plasma and nuclear membranes). In the normal-looking area, the ultrastructural elements in the freshly-formed “dark” neurons were apparently normal; most of them recovered in 1 day; the others were removed from the tissue through an apoptotic-like sequence of ultrastructural changes (the formation of membrane-bound, electrondense, compact cytoplasmic protrusions, and their braking up into membrane-bound, electrondense, compact fragments, which were swallowed by phagocytotic cells). Since these ultrastructural features differ fundamentally from those characteristic of necrosis, it seems logical that, in stark contrast with the prevailing conception, the cause of death of the epilepsy-induced “dark” neurons in the normal-looking cortical area cannot be necrosis. An apoptotic origin can also be precluded by virtue of the absence of its characteristics. As regards the excitotoxic environment, it is assumed that pathobiochemical processes in it superimpose a necrotic-like removal process on already dead “dark” neurons.

Introduction

Ever since the publication of the relevant pioneering paper (Söderfeldt et al., 1983), the “dark” neurons induced by epilepsy have been unanimously believed (Thom et al., 2008) to die through the necrotic pathway. The same has been stated (Auer et al., 2008) for the “dark” neurons induced by ischemia (Smith et al., 1984) or hypoglycemia (Auer et al., 1985). This assumption was based on light- and electron-microscopic observations suggesting that, from necrotic, excitotoxic or contused brain areas, the “dark” neurons produced by these noxae are removed via a necrotic-like sequence of morphological changes.

In contrast, our recent electron-microscopic observations suggested that the mode of death of traumatic (Csordás et al., 2003), electric (Csordás et al., 2003), hypoglycemic (Gallyas et al., 2005) and ischemic (Kövesdi et al., 2007) “dark” neurons is neither necrosis nor apoptosis. This assumption was based on electron-microscopic observations proving that a number of “dark” neurons are produced by these noxae even in non-necrotic, non-excitotoxic or non-contused (apparently normal) tissue areas, from which they are removed via an apoptotic-like sequence of morphological changes.

In the present paper, we investigate whether or not this assumption also applies to “dark” neurons induced by epilepsy. Since there are biochemical differences between the pathological circumstances at issue (Auer and Siesjö, 1988, Liou et al., 2003), it is not evident that the epilepsy-produced “dark” neurons are removed from apparently normal (non-excitotoxic) brain areas via the apoptotic-like sequence of ultrastructural changes, which could support the non-necrotic and non-apoptotic nature of their death.

For the induction of epilepsy, a 4-aminopyridine paradigm was used which produces a relatively large number of “dark” neurons in an apparently normal cortical area not far from a large excitotoxic area (Baracskay et al., in press).