Growth inhibitory effects of a mu opioid on cultured cholinergic neurons from fetal rat ventral forebrain, brainstem, and spinal cord

Cholinergic pathways play a role in respiration in the mammalian brain, and agents that affect respiratory function such as opioid peptides might have positive or negative neurotrophic effects during the development of these cholinergic connections. Rat fetal nerve cell cultures from developmental s tages E14-E18 were established in 96-well plates from ventral forebrain (VF B), an area rich in cholinergic neurons, and from brainstem and rostral spi nal cord, areas where respiratory control systems and cholinergic neurons c o-exist. High affinity H-3-choline uptake was highest in E14 VFB cultures a nd decreased to 20% of this value by E16 and E18. Choline uptakes in E14 br ainstem and spinal cord were only 20% and 13%, respectively, of E14 VFB upt ake. A mu opioid receptor agonist, D-ala(2)-mePhe(4)-gly(ol)(5)]-enkephalin (DAMGO), was tested for its effect on somal area and neurite outgrowth in E16 cultures. Cholinergic neurons were identified by immunostaining with ch oline acetyltransferase antibody. DAMGO (10(-8) M) significantly decreased somal area in VFB cultures and spinal cord, but had no effect on somal area in brainstem. Naltrexone (10(-6) M) reversed this inhibition. Spinal cord cell neurite outgrowth was inhibited by DAMGO, and this inhibition was reve rsed by naltrexone. DAMGO had no significant effect on neurite length in VF B. Brainstem neurite length was paradoxically increased by both DAMGO and n altrexone. It was concluded that mu-selective opioid peptides inhibit growt h of cultured cholinergic neurons in VFB and spinal cord, but not in the br ainstem. There was no evidence for endogenous opioid activity in either VFB or spinal cord cultures. (C) 1999 Elsevier Science B.V. All rights reserve d.