METALLOTHIONEIN INDUCTION IN NEONATAL RAT PRIMARY ASTROCYTE CULTURES PROTECTS AGAINST METHYLMERCURY CYTOTOXICITY

Metallothionein (MT) protein and mRNA levels were monitored following exposure of rat neonatal primary astrocyte cultures to methylmercury ( MeHg). MT-I and MT-II mRNAs were probed on northern blots with an [alp ha-P-32]dCTP-labeled synthetic cDNA probe specific for rat MT mRNA. MT -I and MT-II mRNAs were detected in untreated cells, suggesting consti tutive MT expression in these cells. The probes hybridize to a single mRNA with a size appropriate for MT, similar to 550 and 350 bp for MT- I and MT-II, respectively. Expression of MT-I and MT-II mRNA in astroc yte monolayers exposed to 2 x 10(-6) M MeHg for 6 h was increased over MT-I and MT-II mRNA levels in controls. Western blot analysis reveale d a time-dependent increase in MT protein synthesis through 96 h of ex posure to MeHg. Consistent with the constitutive expression of MTs at both the mRNA lever and the protein level, we have also demonstrated a time-dependent increase in MT immunoreactivity in astrocytes exposed to MeHg. The cytotoxic effects of MeHg were measured by the rate of as trocytic D-[H-3]aspartate uptake. Preexposure of astrocytes to CdCl2, a potent inducer of MTs, completely reversed the inhibitory effect of MeHg on D-[H-3]aspartate uptake that occurs in MeHg-treated astrocytes with constitutive MT levels. Associated with CdCl2, treatment was a t ime-dependent increase in astrocytic MT levels. in summary, astrocytes constitutively express MTs; treatment with MeHg increases astrocytic MT expression, and increased MT levels (by means of CdCl2 pretreatment ) attenuate MeHg-induced toxicity. increased MT expression may represe nt a generalized response to heavy metal exposure, thus protecting ast rocytes and perhaps also, indirectly, juxtaposed neurons from the neur otoxic effects of heavy metals.