Calcium/calmodulin-dependent protein kinase II clusters in adult rat hippocampal slices

Abstract

We have previously reported the formation of calcium/calmodulin-dependent protein kinase II (CaMKII) clusters approximately 110 nm in diameter in hippocampal neurons in culture and in the intact adult brain, under conditions that simulate ischemic stress and increase [Ca(2+)](i) [Dosemeci et al. (2000) J. Neurosci. 20, 3076-3084; Tao-Cheng et al. (2001) Neuroscience 106, 69-78]. These observations suggest that ischemia-like conditions that prevail during the dissection of brain tissue for the preparation of hippocampal slices could lead to the formation of CaMKII clusters. We now show by pre-embedding immuno-electron microscopy that, indeed, CaMKII clusters are present in the CA1 pyramidal neurons in hippocampal slices from adult rats fixed immediately after dissection, and that the number of CaMKII clusters increases with the delay time between decapitation and fixation. Moreover, CaMKII clusters are typically localized near the endoplasmic reticulum. When acute slices are allowed to recover in oxygenated medium for 2 h, CaMKII clusters mostly disappear, indicating that clustering is reversible. Also, the postsynaptic density, another site for CaMKII accumulation under excitatory conditions, becomes thinner upon recovery. Treatment of recovered slices with high potassium for 90 s causes the re-appearance of CaMKII clusters in nearly all CA1 pyramidal cells examined. On the other hand, when dissociated hippocampal neurons in primary culture are exposed to the same depolarizin…