Acrylamide disturbs the subcellular distribution of GABAA receptor in brain neurons.

Mechanisms underlying the action of acrylamide on neurons were studied by monitoring the expression of GABA(A) receptor (R) in cultured brain neurons derived from chicken embryos. In situ trypsinization of the neurons and 3H-flunitrazepam binding assay were employed to examine the subcellular distribution of GABA(A)R. A 3-h exposure of the cultured neurons to 10 mM of acrylamide raised reversibly the proportion of intracellular (trypsin-resistant) 3H-flunitrazepam binding sites by about 48% and decreased cell surface binding 24% from respective control values, without altering total cellular binding and the affinity of the ligand. Moreover, the acrylamide treatment induced more intense perikaryal immunostaining of GABA(A)R alpha subunit proteins than that in control neurons but did not change the total level of cellular alpha immunostain, in accordance with the binding data. In the cell bodies of acrylamide-treated neurons, the level of neurofilament-200 kDa proteins was similar to control, whereas the tubulin protein content was significantly lowered approximately 51% from control, as revealed by quantifying the immunostained cytoskeletal elements. In addition, electron microscopic observations found reductions in the numbers of microtubules and neurofilaments in the perikarya of acrylamide-treated neurons. As exhibited by the 3H-leucine and 3H-monosaccharide incorporation experiments, the exposure to acrylamide inhibited the rate of general protein synthesis in the culture by 21%, while the rate of glycosylation remained unaltered. Furthermore, in situ hybridization analysis showed that acrylamide did not modify the expression of GABA(A)R alpha subunit mRNAs. Taken together, these data suggest that acrylamide may downregulate the microtubular system and disintegrate neurofilaments, and thereby block the intracellular transport of GABA(A)R, resulting in the accumulation of intracellular receptors. Copyright 2002 Wiley-Liss, Inc.