The ability of the nervous system to generate coherent behavior is dependent on a vast number of precise synaptic connections between neurons. Very little is known about the molecular mechanisms which generate these connections, but all tenable hypotheses ultimately postulate the existence of molecules, differing in kind or quantity from cell to cell, which mediate the necessary recognition. We are using the leech embryo as a developmental system, where we can relate chemical specificity to unique connectivity. Certain leech sensory afferent neurons are identifiable by their carbohydrate epitopes. The mannose-containing Lan3-2 epitope is common identifiable by their carbohydrate epitopes (the Laz2-369, Laz7-79 and 141 epitopes) are expressed by different subpopulations of sensory afferents: the putative wave-, chemo-, and heatdetectors. The Lan3-1 epitope has been shown to be important in the process of defasciculation of sensory afferents as they reach the synaptic neuropile, a process which allows the sensory afferents to project their appropriate target domains. The Lan3-2 mediated mannose-specific recognition also appears to be important in maintaining continued occupancy of these domains. Here we propose to further investigate the developmental significance of carbohydrate recognition mediated by four different carbohydrate epitopes. Using FAb fragments and neoglycoproteins, we will experimentally manipulate the newly formed monosynaptic connections between sensory afferents and their identified postsynaptic partners, e.g. an interneuron of the local bending reflex, cell 202. Alteration in the apposition of sensory afferent axonal arbors with the neuritic domains of their postsynaptic neurons will be quantified in laser scanned horizontal sections using confocal microscopy. Alteration in the monosynaptic connections between sensory afferents and their postsynaptic neurons will be measured physiologically, by both electrophysiologically recording extracellular spike activity and by measuring changes in Ca++ flux through voltage-gated Ca++ channels. Accumulating evidence shows that functional classes of neurons express selective carbohydrate epitopes on their cell surfaces throughout phylogeny. Manipulating carbohydrate recognition mechanisms in the molecularly accessible simple leech nervous system therefore promises to elucidate general biological principles underlying the formation of neuronal connectivity relevant to development and regeneration.
Showing the most recent 10 out of 22 publications
