Stereological Analysis of the Motor Cortex and the Hippocampus of the Dermatan-4O-sulfotransferase1 (Chondroitin Sulfotransferase 14) knockout mice

Abstract

Chondroitin/dermatan sulfate proteoglycans are major components of the extracellular matrix important for cell migration and axonal pathfinding. The different chondroitin/dermatan sulfate structures are generated by sulfation and epimerisation of glucuronic/iduronic acid. In order to distinguish between functions of chondroitin and dermatan sulfates, a mouse deficient in dermatan-4O-sulfotransferase1 (Chst14 -/-), the key enzyme indispensable for the formation of iduronic acid-containing blocks in dermatan sulfates, has been recently created. In order to uncover the roles of dermatan sulfates in cortical development, I studied these Chst14 -/- mice with respect to morphological abnormalities in the motor cortex and hippocampus, well-defined brain structures known to be functionally relevant for higher brain functions including learning and memory. Morphometric as well as stereological analyses of immunohistochemically identified major cell types (neurons, neuronal subpopulations, astrocytes, oligodendrocytes and microglia) were performed on 2-month-old Chst14 -/- mice and wild-type (Chst14 +/+) littermates. Analyses of gross anatomical variables revealed that Chst14 -/- mice had significantly reduced body weight (-21%) compared with Chst14 +/+ littermates, which could be explained by reduction in the mass of bones, heart, liver, kidneys and other organs. In contrast, the Chst14 -/- brains were normal in size and had no gross morphological abnormalities. The cortical thickness in the motor cortical area as well as the total area of the hippocampus and its subfields were also not altered in Chst14 -/- mice. However, the pyramidal cell layer and stratum radiatum in CA3 were reduced in size by 13% and 16%, respectively, in Chst14 -/- compared with Chst14 +/+ mice. Additionally, the numerical density of pyramidal cells was reduced by 24% and density of parvalbumin-positive (PV+) interneurons was increased by 26% in the CA3 region of Chst14 -/- mice. Since PV+/principal cell ratio in the CA3 of Chst14 -/- was by 76% higher than in Chst14 +/+ mice, I also analyzed densities of perisomatic inhibitory terminals on pyramidal cells using confocal microscopy. While in the CA1 region there was no difference between the genotypes, the terminal density was reduced by 13% in the CA3 of Chst14 -/- compared with Chst14 +/+ mice. Other regions of the hippocampus as well as the motor cortex were not affected by the mutation. Major cell populations, including total cell numbers and NeuN+ neurons in the cortex, principal cells in the hippocampus, PV+ and reelin+ interneurons, CNPase+ oligodendrocytes and S100+ astrocytes had similar densities in the two genotypes. One notable exception was Iba1+ microglia, which was increased in density in all studied areas (motor cortex +26%; CA1 +36%; CA3 +34%; DG +22%). This study gives important first insights into the structural changes in the brain caused by Chst14 deletion. Further electrophysiological and behavioural experiments are needed to understand the functional consequences of the observed structural aberrations and the role of Chst14 and dermatan sulfates in neural development and plasticity.