Regeneration of central and peripheral synaptic connections in the locomotor system of the pteropod molluscClione limacina

Abstract

The neural network underlying rhythmic wing movements in the mollusc CIione limacina is well-studied. Two different groups of motoneurons innervate two distinct groups of wing muscles. The locomotor rhythm generated in the left and right pedal ganglia is synchronized by interneurons. When the axons of the locomotor motoneurons are crushed, numerous fine neurites sprout towards the denervated muscles and reach them in 8-15 days. At this stage motoneurons project to and synapse on not only correct but equally incorrect muscle targets. After 2 weeks of regeneration the number of incorrect neurites and synaptic connections begins to decrease and following 1.5-2 months all incorrect connections are eliminated, incorrect axons are withdrawn and the behavioral deficit is compensated. In this study the regeneration of intemeurons and the growth profiles of inter-and motoneurons were also studied in vitro. Two individually isolated pedal ganglia were co-cultured in three different configurations: a) the wing nerve stump from one ganglion was fixed against the commissural stump from another ganglion; b) the wing nerve stumps were fixed against each other; c) the commissural stumps were fixed against each other. Under the above experimental conditions we found that the interneurons were able to cross only the contact between two commissural stumps, and in this case found their original targets, restored correct connections and synchronized the rhythm in two pedal ganglia. In contrast, motoneurons were able to cross all types of contacts.

Projection pattern and target selection of Clione limacina motoneurons sprouting within an intact environment

“…In a previous study (Panchin et al, 1998) where only a wing nerve was cut in the whole animal, the neurite growth toward the contralateral side was weak, and most neurites grew out of the ganglion to the ipsilateral peripheral nerves. To facilitate the growth of new sprouts to the contralateral ganglion and wing, all exits from one of the pedal ganglia were severed (see Materials and Methods).…”

“…A CNS-"split wing" preparation was made ( Fig. 1C; Panchin et al, 1998). The posterior edge of the wing contralateral to the denervated ganglion was cut with scissors and the two layers of swimming muscles were separated ( Fig.…”

“…In order to choose among these possibilities, we used the Clione neuromuscular regeneration model to study the growth of novel neurites of wing motoneurons into the intact contralateral wing nerve. If only the wing nerve was cut in the whole animal, the majority of newly formed neurites grew within the ipsilateral part of the CNS, rarely invading the contralateral pedal ganglion (Panchin et al, 1998). To enhance the growth towards the contralateral part of the CNS and into the contralateral wing nerve, we deprived the growing neurites of all possible tracts except for the tract to the contralateral pedal ganglion.…”

“…Later, the incorrect synapses and neurites are withdrawn. This leads to restoration of the correct morphology of motoneurons and correct muscle innervation (Panchin et al, 1998). Additional data on the pathfinding and target selection of the sprouting motoneurons during wing reinnervation were obtained in the experiments on regenerating animals and during in vitro culturing of muscles and ganglia.…”

“…Each secondary branch projects to only one of the two muscle layers and carries axons of only one of the two classes of motoneurons (either only D or only V; Fig. 1C; Panchin et al, 1998).…”