Metallothionein induction in fetal rat brain and neonatal primary astrocyte cultures by in utero exposure to elemental mercury vapor (Hg 0)

Abstract

Brain metallothionein (MT) protein and mRNA levels were determined in the fetal rat following in utero (gestational days 7–21) exposure to elemental mercury vapor (Hg 0; 300 μg Hg/m 3; 4 h/day). Total RNA was probed on northern blots with [ α- 32P]dCTP-labeled synthetic cDNA probes specific for rat MT isoform mRNAs. The probes for MT-I and MT-II mRNA hybridized to a single band of approximately 550 and 450 nucleotides, respectively. Expression of whole brain MT-I mRNA in full-term fetal rats (day 21) was significantly increased ( P<0.03) by in utero exposure to Hg 0 compared to nonexposed controls. This corresponded to a 14-fold increase ( P<0.001) in fetal brain Hg concentration after in utero Hg 0 exposure. In addition, astrocytes from both control and in utero Hg 0-exposed fetuses were isolated, and neonatal primary astrocyte cultures were established and maintained in vitro for up to 3 weeks without additional experimental intervention. Astrocyte monolayers derived from in utero Hg 0-exposed fetuses consistently expressed increased abundance of MT-I mRNA transcripts after 1, 2, and 3 weeks in culture ( P<0.03, P<0.01, and P<0.03, respectively) compared with controls. The abundance of astrocyte MT-II mRNA was unchanged at 1 and 2 weeks in culture, but was significantly increased at 3 weeks in cultures derived from brains of Hg 0-exposed fetuses ( P<0.04). Consistent with the increase in MT mRNA, an increase in astrocytic levels of MT proteins was noted by western blot analysis and MT-immunoreactivity. These studies suggest that in utero exposure to Hg 0 induces brain MT gene expression, and that MT mRNAs and their respective proteins are useful quantitative biochemical markers of intrauterine exposure to Hg 0, a potentially cytotoxic challenge to astrocytes in the developing brain. It is concluded that induction of MT by fetal/neonatal astrocytes represents an attempt by these glial cells to protect against Hg cytotoxicity in maintaining cerebral homeostasis.