Organotypic slice cultures are being used increasingly to understand the physiology and molecular basis of synaptic plasticity (Gèhwiler et al. 1997). Here we describe the developmental profile of cytoskeletal and synaptic marker proteins: synaptophysin (a pre-synaptic vesicle-associated protein), PSD95 (a post-synaptic protein found clustered with NMDA receptor subunits), β-tubulin (a microtubule stabilisation and growth GTPase putatively linked to NMDA receptors via PSD95) and a brain-specific neuroplastin np65 that is involved in hippocampal synaptic plasticity (Smalla et al. 2000).

The developmental expression of these proteins was compared in rat hippocampal acute and organotypic slices using Western blot analysis. Acute hippocampal slices were taken from humanely killed post-natal day (P)5 to adult P120 rats. Organotypic hippocampal slices, prepared from P7 rats (Stoppini et al. 1991), were taken between 0 and 21 days in vitro (DIV). Organotypic slices were cultured in control conditions or the presence of the NMDA- and/or AMPA-receptor antagonists: AP5 (100 µM) and CNQX (20 µM), respectively, for 7 DIV.

Expression of np65 in organotypic slices mimicked that in acute slices, peaking between P21-28 and 14-21 DIV. Synaptophysin and PSD95 displayed very similar developmental profiles in acute and organotypic slices rising in expression from P14 and 7 DIV onwards. β-Tubulin displayed decreasing post-natal expression in both acute and organotypic slices.

Chronic treatment of slice cultures with glutamate receptor antagonists did not affect the total expression of np65 compared with control. AP5 alone had no effect upon PSD95, synaptophysin and β-tubulin levels, although CNQX alone decreased PSD95 expression by ca 60 % compared with control. A synergistic effect of CNQX and AP5 together further reduced PSD95 levels to ca 90 % and also decreased the expression of synaptophysin and β-tubulin, both ca 40 % compared with control.

Our results indicate that organotypic slice cultures can be used as a reliable model to investigate functional and molecular changes in synaptogenesis and synapse stability. Furthermore chronic treatment with AMPA- and NMDA-receptor antagonists reduced the expression of PSD95, synaptophysin and β-tubulin suggesting a perturbation of synaptic function.

We acknowledge the support of the BBSRC (111/NEU15396).