Calbindin D28K expression in relation to granule cell dispersion, mossy fibre sprouting and memory impairment in hippocampal sclerosis: A surgical and post mortem series

Surgical cases labelled with calbindin (CB) (top row), NeuN (middle row) and dynorphin (bottom row). The three patterns of calbindin expression (which formed the three groups) are shown. (a) Pattern 1: The basal granule cells are predominantly CB negative and the outermost granule cells, dispersed in the molecular layer are CB positive (inset shows very occasional CB positive cell with basal axon: arrowhead). (b) Pattern 2: The majority of granule cells are positive with strong labelling of the apical dendrites in the molecular layer and in CA4 mossy fibres (normal pattern). (c) Pattern 3: A virtual absence of CB expression in the granule cells. (d) NeuN showing a bilaminar pattern of granule cell dispersion with a cell free zone between the basal and most migrated granule cells (bGCD), (e) granule cell dispersion (GCD) and (f) no granule cell dispersion with a compact cell layer. (g) Dynorphin with Grade 2 mossy fibre sprouting with a broad band of axonal labelling in the molecular layer, (h) Grade 1 mossy fibre sprouting with more patchy labelling in the inner molecular layer and (i) Grade 0 no mossy fibre sprouting with positivity for dynorphin only in CA4. Bar=35μm.

Bar chart showing the mean distances of CB-positive and CB-negative granule cells in relation to the basal cell layer. Over all cases the distances were significantly greater for the positive cells (p<0.01) but between the groups this finding remained significant for Group 1 only (p<0.001). (Numbers in brackets indicate the number of cases in each group in which measurements were made; significant measurement indicated with *.)

A surgical case with hippocampal sclerosis from Group 1. (a) Calbindin and dynorphin (c) at low magnification show similar distribution of labelling in the molecular layer and the CA4/3 region (arrows). Pattern 1 of calbindin expression (b) and mossy fibre sprouting (d) is confirmed at high magnification. (e) Confocal microscopy confirms a lack of co-localisation in the molecular layer with calbindin expressed in cell bodies and apical dendrites of granule cells and dynorphin in the sprouted mossy fibres. In addition there are less dynorphin positive sprouted fibres beyond the outermost calbindin positive cells (arrow head) (f) but there is co-localisation between dynorphin and calbindin in residual CA4 mossy fibres (arrow). Bar in a and c=10μm, in b, d and f=35μm and in e=50μm.

Post mortem cases. (a) Left and (b) right hippocampus from control cases with normal calbindin expression. Unilateral hippocampal sclerosis (case EP200 in Table 2) showing asymmetry of calbindin expression with a normal pattern observed on the side without sclerosis (c) and absence of staining in the contralteral sclerosed hippocampus (d). This pattern of calbindin was retained in all blocks into the hippocampal tail (insets in c and d). (e) In post mortem cases relatively restricted calbindin expression, present mainly in the outermost granule cells, was also observed. Bars in a–d=10μm and e=25μm.