Production of sweat gland cholinergic differentiation factor depends on innervation

Sympathetic neurons innervating sweat glands undergo a target-directed developmental switch in neurotransmitter properties. Using cultured sympathetic neurons as a bioassay for cholinergic differentiation factors, we and others found that extracts containing soluble proteins from developing and adult footpads caused the same changes in transmitter properties in sympathetic neurons in vitro that the target does in vivo. In the present studies, using footpads from Tabby mutant mice that lack sweat glands, we found that the presence of sweat glands is correlated with the presence of cholinergic differentiation activity in footpad extracts. We examined the conditions necessary for secretion of differentiation activity from primary cultures of sweat gland cells. Surprisingly, sweat gland cells cultured alone do not produce or secrete cholinergic differentiation activity. When grown in the presence of sympathetic neurons, however, gland cells induce cholinergic function, increase vasoactive intestinal peptide content, and reduce catecholamine production in the neurons. Medium conditioned by sweat gland/neuron cocultures has a similar effect on the transmitter properties of cultured sympathetic neurons, indicating that the target influence on phenotype is mediated by a secreted factor(s). The innervation-dependence of cholinergic differentiation factor production provides evidence that reciprocal interactions between neurons and sweat glands are necessary for acquisition of the mature transmitter phenotype.