Acetylcholine analogue stimulates DNA synthesis in brain-derived cells via specific muscarinic receptor subtypes

LITTLE is known about the factors which regulate the growth and development of the mammalian brain. Although proliferation of neuronal cells ceases relatively early in development, certain types of glial cells proliferate and differentiate mainly perinatally1. In the perinatal period, the ability of acetylcholine to stimulate phosphoinositide (PI) hydrolysis in brain reaches peak levels2, and indeed the stable acetylcholine analogue carbachol can stimulate PI hydrolysis of primary neonatal astroglial cells3. As PI hydrolysis is thought to be important in the regulation of cell proliferation4-6, we investigated whether cellular DNA synthesis can be induced by carbachol. Our results show that carbachol stimulates DNA synthesis via muscarinic acetylcholine receptors (mAChRs), in primary astrocytes derived from perinatal rat brain, in an age-dependent fashion. Carbachol is also mitogenic in certain brain-derived astrocytoma and neuroblastoma cell lines, as well as in Chinese hamster ovary (CHO) cells expressing recombinant muscarinic receptors. DNA synthesis is strongly activated by car-bachol in those brain-derived cell lines and transfected CHO cells that express mAChR subtypes which activate PI hydrolysis efficiently, and poorly activated in cells expressing mAChR sub-types which only weakly activate PI hydrolysis. These results strongly support a role for acetylcholine in regulating astroglial cell growth in the developing brain, and indicate that the specificity of acetylcholine-induced cell proliferation may be determined by the expression of those mAChR subtypes which activate PI hydro-lysis.