THE MODULATORY CHOLINERGIC SYSTEM IN GOLDFISH TECTUM MAY BE NECESSARYFOR RETINOTOPIC SHARPENING

The cholinergic circuit within the tectum and the cholinergic input fr om the nucleus isthmi mediate a presynaptic augmentation of retinotect al transmitter release via nicotinic receptors. In this study, the cho linergic systems were either eliminated using the cholinergic neurotox in AF64A or blocked using nicotinic antagonists to test for effects on the activity-driven sharpening of the regenerating retinotectal proje ction. The effectiveness of the AF64A was verified by recording field potentials elicited by optic tract stimulation and by immunohistochemi cal staining for choline acetyltransferase (ChAT). At 1 week after int racranial (IC) injection of AF64A (12 to 144 nmoles) into the fluid ab ove the tectum, field potentials showed a selective dose-dependent dec rement of the cholinergic polysynaptic component with no effect on the amplitude of the glutamatergic monosynaptic component. The decrement was only partially recovered in recordings at 2 and 6 weeks. In normal fish, the ChAT antibody stains a population of periventricular neuron s, their apical dendrites, and a dense plexus within the optic termina l lamina that consists of their local axons and fine dendrites and of input fibers from the nucleus isthmi. One week after IC AF64A injectio n (48-72 nmoles), most immunostaining in superficial tectum was lost b ut most neuronal somas in the deep tectum could still be seen, and sta ining in the tegmentum below the tectum was completely intact. At 2 we eks and later, the staining of neuronal somata largely recovered, but staining of the superficial plexus did not. AF64A treatment at 18 days after nerve crush, when regenerating retinal fibers are beginning to form synapses, prevented retinotopic sharpening of the projection. Rec ordings showed a rough retinotopic map on the tectum but the multiunit receptive fields (MURFs) at each tectal point averaged 34 deg vs. 11 deg in vehicle-injected control regenerates. AF64A treatment before ne rve crush also blocked sharpening, ruling out a direct effect on retin al growth cones or retinal fibers, as AF64A rapidly decomposes, wherea s its effect on the cholinergic fibers is long-lasting. IC injection o r minipump infusion of the nicotinic antagonists alpha-bungarotoxin (a lpha BTX), neuronal bungarotoxin (nBTX), and pancuronium during regene ration also prevented sharpening (MURFs averaging 29.4 deg, 33.0 deg, and 31.4 deg, respectively). Control Ringer's solution infusions or in jections over the same period (19-37 days postcrush) had no effect on regenerated MURF size (11.7 deg). The results show that the cholinergi c innervation, which modulates transmitter release, is required for ac tivity-driven retinotopic sharpening, thought to be triggered by NMDA receptor activation.