Temporal Lobe Epilepsy Neuropathology and the Dentate Gyrus
One of the major concepts that dominated the epilepsy research community in 1980s was the importance of underlying neuropathology in the hippocampus. For the purposes of this review, the hippocampus is defined as areas CA1, CA2, CA3, and the dentate gyrus (DG). There was a common view that temporal lobe epilepsy (TLE) was characterized by a pattern of hippocampal neuronal loss that was originally called Ammon’s horn or hippocampal sclerosis and then, as neuronal loss in extrahippocampal areas became appreciated, mesial temporal sclerosis (MTS).1 In the hippocampus, neuronal loss mainly included the DG hilus and pyramidal cell layers of areas CA1 and CA3. The DG and area CA2 cell layers were relatively resistant.1
Notably, it was often assumed that MTS led to epilepsy. One argument was the finding that seizures in patients seemed to start in the sclerotic hippocampus.2 Surgical removal often helped, supporting the hypothesis, but did not always stop clinical seizures. How the hippocampus could be important but its removal not provide a cure could be explained by the concept of secondary epileptogenesis,3 where secondary foci assume the role of the primary focus if the primary focus is removed. Still, explaining hippocampal neuropathology continued to attract attention.
At the core of the issue is selective vulnerability, that is, why some neurons are vulnerable relative to others. Experiments began to focus on characteristics of individual hippocampal neurons to understand this issue, made possible by the increasing acceptance of the hippocampal slice preparation and with it, the ability to make stable intracellular recordings. Using improved markers of single cells such as biocytin, the basic neuronal properties and morphologies of hippocampal neurons were clarified, and an increasingly detailed map of their circuitry emerged. This understanding proceeded in parallel with cellular investigations of induced seizure-like activity in slices of normal animals, leading to predictions about how the characteristics of the cells and circuitry contributed to seizures. The techniques have now been superseded by more advanced imaging and recording methods, including recordings in awake head-fixed animals, as well as viral-based strategies and techniques to record simultaneously from many areas rather than one cell. Therefore, the understanding of cells and their circuitry continues, but the “neurocentric” view has given way to one that includes glia, the vasculature, and immune cells.
Most studies were made of CA1 and CA3 initially because the DG didn’t survive well after the slice procedure. Better equipment to section the brain and methods to limit damage during the slice procedure helped. In the DG, the relatively resistant granule cells (GCs) lay juxtaposed to vulnerable hilar neurons, making the DG attractive to understand reasons for selective vulnerability. At first GCs and hilar neurons were characterized. Hilar neurons became divided into 2 categories: the glutamatergic mossy cells (MCs) and diverse GABAergic neurons.4 Vulnerable hilar neurons were identified as the MCs and a subset of hilar GABAergic neurons that expressed the neuropeptide somatostatin. Later the DG GABAergic neurons were categorized by their axonal targets,5 and despite the developments of other classifications6 the nomenclature based on the axon has been widely adopted. Hilar somatostatin-expressing cells are now often referred to as the hilar cells with a terminal projection associated with the perforant pathway (the outer 2/3 of the DG molecular layer), or HIPP cells.
A common hypothesis in the 1980s to 1990s for MCs and HIPP cell vulnerability was based on the research being done at the time about excitotoxicity, and the common view that TLE was caused by a brain insult or injury. Based on these views, it seemed logical that neuronal death after brain insults is due to excitotoxicity induced by the brain insult.7 A critical step in excitotoxicity was the escalation of intracellular calcium levels during strong glutamatergic stimulation. The role of glutamate and calcium in excitotoxicity arose from experiments often in areas or systems besides hippocampus8 and the role of glutamatergic afferents to the DG arose from the repetitive stimulation animal model.9,10 In this model, afferent stimulation was triggered repeated in an anesthetized rat using a stimulating electrode placed into the perforant path, the major afferent to the DG. Neuroanatomical studies showed that vulnerable MCs and HIPP cells died after the stimulation, but not the DG GCs.11 It was surprising that the primary type of GABAergic cell survived (called basket cells; now called perisomatic-targeting cells, often expressing parvalbumin (PV)) because it was often assumed that they were vulnerable, and deficient GABAergic inhibition was critical in seizure generation.12
Using antibodies to label different cell types, studies showed that there was a low expression of calcium binding proteins (calbindinD28K (CaBP), PV) in the MCs and HIPP cells but high expression of CaBP in the GCs and PV in the perisomatic-targeting cells.13 Later it was shown that chelating intracellular calcium in MCs and other hilar interneurons could protect them in a slice model of repetitive stimulation.14 However, MCs and HIPP cells lack other proteins that GCs have (eg striatal-enriched protein tyrosine phosphatase; STEP), and new molecular techniques suggest that STEP is important for neuroprotection.15 The “calcium-binding protein hypothesis” is now discussed much less because, in part, selective vulnerability in other brain areas is not necessarily correlated with calcium binding protein content.16 The field has also become more complex with diverse roles and regulation of intracellular calcium and new interest in apoptosis of hilar cells.17
Other hypotheses arose to address selective vulnerability based on cellular and circuit properties, perhaps because of the wealth of new data arising from new improved methods such a patching visualized neurons. For example, GCs had resting membrane potentials (RMPs) that were hyperpolarized compared to most other neurons in the hippocampus.18,19 So called “tonic” inhibition and other aspects of the powerful GABAergic inhibition of GC firing emerged20 as well as complex regulation of Gamma-amino-butyric acid (GABA) receptors.21,22 The GCs also express ion channels which make them stop firing if a persistent depolarizing input occurs, called spike frequency adaptation.18,19 Vulnerable MCs and HIPP cells showed more depolarized RMPs than GCs, and less adaptation, although the resistant perisomatic-targeting cells also had depolarized RMPs and little adaptation.23-25 There was an increasingly common view that the normal “quiescence” of GCs, rarely firing action potentials, explained the GC resistance to brain insults. Without as much action potential discharge, it was suggested that excitotoxicity would be unlikely. The relative quiescence of GCs was later supported by additional recordings in vivo26 and is now considered a dogma. It appears critical to normal cognitive functions dependent on the DG. Conversely, MCs are highly active both in vitro and in vivo,24,27 and work has suggested that the normally high activity in MCs is also important for cognitive functions related to the DG.28 Thus, what is good for normal functions seems to place the brain at risk for MTS-like pathology and possibly TLE.
Hypotheses to explain selective vulnerability are still an active area of research, in part because it is as important as ever to address the debilitating effects of brain injury. On the other hand, neuroprotective strategies have not been a “magic bullet” in preclinical epilepsy research.29 This has led to new energy into other pressing questions. For example, the way genetics applies to epilepsy has become an area of increasing attention. One reason is that studies of “idiopathic” epilepsy have revealed the importance of genetics.30,31 Methodological advances have allowed both clinical and basic scientists to increasingly ask questions related to genes.