A Delayed Role for Nitric Oxide-Sensitive Guanylate Cyclases in a Migratory Population of Embryonic Neurons

Neuronal differentiation requires a coordinated intracellular response to diverse extracellular stimuli, but the role of specific signaling mechanisms in regulating this process is still poorly understood. Soluble guanylate cyclases (sGCs), which can be stimulated by diffusible free radical gasses such as nitric oxide (NO) and carbon monoxide (CO) to produce the intracellular messenger cGMP, have recently been found to be expressed within a variety of embryonic neurons and implicated in the control of both neuronal motility and differentiation. Using the enteric nervous system (ENS) of the moth,Manduca sexta,we examined the role of NO and NO-sensitive sGCs during the migration and differentiation of an identified set of migratory neurons (the EP cells). Shortly after the onset of their migration, a subset of EP cells began to express NO-sensitive sGC activity (visualized with an anti-cGMP antiserum). Unlike many neurons in the central nervous system, the expression of sGC activity in the EP cells was not transient but persisted throughout subsequent periods of axon elongation and terminal branch formation on the gut musculature. In contrast, nitric oxide synthase activity (visualized using NADPH–diaphorase histochemistry) was undetectable in the vicinity of the EP cells until the period of synapse formation. Manipulations designed to alter sGC and NOS activity in anin vivoembryonic culture preparation had no discernible effect on either the migration or axonal outgrowth of the EP cells. In contrast, inhibition of both of these enzymes resulted in a significant reduction in terminal synaptic branch formation within the postmigratory neurons. These results indicate that while NO-sensitive sGC activity is expressed precociously within the EP cells during their initial migratory dispersal, a role for this signaling pathway can only be demonstrated well after migration is complete, coincident with the formation of mature synaptic connections.