The method of isolation of the crayfish abdominal stretch receptor maintaining a connection of the sensory neuron to the ventral nerve cord ganglion.

Abstract

The crayfish stretch receptor consisting of the single mechanoreceptor neurons enveloped by satellite glial cells is the simplest functioning neuroglial preparation. However, during isolation, its axons are usually transected that eliminates afferent regulation and induces complex axotomy-related signaling responses in neurons and satellite glia. We developed new microsurgical method of crayfish stretch receptor isolation, which preserves connections of sensory neurons to the ventral nerve cord ganglion. The stretch receptor may either remain on the abdominal carapace, or be completely isolated. In both cases, it may be either intact, or axotomized. The integrity of axons was confirmed by firing recording from proximal and distal axon points. Normal, necrotic and apoptotic cells were visualized using double fluorochroming with Hoechst 33342 and propidium iodide. The isolated mechanoreceptor neurons maintain regular firing during 8-10 or more hours. Glial cells surrounding non-axotomized neurons demonstrate lower necrosis and apoptosis levels than the axotomized ones. Unlike the existing method, in which the sensory neurons were axotomized, the present method preserves links between the sensory neurons and the ganglion and makes possible to avoid consequences of axotomy in neurons and satellite glia. The present neuroglial preparation may be used as a simple but informative model object in studies of axotomy-induced degeneration and survival of peripheral neurons, the role of glia in neuron injury, the signaling mechanisms of neuroglial interactions, and the effects of diverse physical and chemical factors on neuronal and glial cells.