Calcium influx-independent depression of transmitter release by 5-HT at lamprey spinal cord synapses

Insight into the mechanisms of synaptic modulation is important for understanding information processing in the central nervous system (CNS). Modulation of synaptic transmission occurs at several sites: at presynaptic terminals, by the activation of presynaptic receptors; at postsynaptic cells, by change in the postsynaptic receptor properties; and at the synaptic cleft, by change in the clearance rate of released transmitter. At the presynaptic terminal there exist at least four identifiable targets for modulation of release by receptors. (1) Changes in presynaptic resting Ca2+ concentrations, either by the opening or closing of presynaptic ligand-gated Ca2+ channels (Berretta & Jones, 1996; Schwartz & Alford, 1998; Cochilla & Alford, 1999; Glitsch & Marty, 1999) or by the regulation of Ca2+ release from internal stores (Cochilla & Alford, 1999). (2) Mechanisms that alter Ca2+ channel gating may also play a role (Dunlap & Fischbach, 1978; Takahashi et al. 1996; Wu & Saggau, 1997). (3) Changes in the properties of K+ (Ponce et al. 1996; Saugstad et al. 1996) or Na+ (Ma et al. 1997) currents will alter voltage-gated Ca2+ entry during action potentials. (4) G-protein-coupled receptor-mediated mechanisms acting on the release machinery ‘downstream’ of Ca2+ entry have been implicated in the modulation of transmitter release (Hilfiker & Augustine, 1999). These downstream mechanisms would not require any modulation of Ca2+ entry into the terminal, unlike other mechanisms suggested above (Silinsky, 1984; Scholz & Miller, 1992; Gereau & Conn, 1995; Scanziani et al. 1995). The activation of a G-protein subunit (Kowluru et al. 1996; Zhang et al. 1998) may directly alter vesicle fusion to the presynaptic membrane. Indeed Gβγ has recently been shown to bind directly to syntaxin (Jarvis et al. 2000). Alternatively, G-protein subunit activation may initiate other signal transduction cascades (e.g. by activating protein kinase C and Ca2+-calmodulin-dependent kinase II; Pasinelli et al. 1995). 5-Hydroxytryptamine (5-HT) acts throughout the CNS, at presynaptic and postsynaptic targets (Wallen et al. 1989; Hounsgaard & Kiehn, 1989; Buchanan & Grillner, 1991). Like other G-protein-coupled receptors, a number of mechanisms have been proposed for the presynaptic action of 5-HT receptors in altering transmitter release. These include the inhibition of Ca2+ channels (El Manira et al. 1997) or enhancement (Dale & Kandel, 1990; Delaney et al. 1991) or inhibition (Shupliakov et al. 1995) of release downstream of Ca2+ entry to the terminal. 5-HT1A, 5-HT1B and 5-HT1D receptors have been demonstrated to depress glutamate release in rat amyglada (Cheng et al. 1998), in the entorhinal cortex (Schmitz et al. 1998) and in the hippocampus (Schmitz et al. 1995). 5-HT-mediated depression of glutamatergic inputs to spinal motor neurons has been observed across many vertebrate groups from cyclostomes (Buchanan & Grillner, 1991) to mammals (Singer & Berger, 1996). Access for recording techniques to the presynaptic terminal is necessary to elucidate the cellular mechanisms involved in presynaptic modulation. However, the small size of vertebrate presynaptic terminals usually prevents a direct study of these mechanisms. In the lamprey spinal cord, reticulospinal axons make en passant excitatory synapses onto motor neurons and ventral horn interneurons. The large unmyelinated axons (diameter up to 100 μm) allow an easy access for both direct electrophysiological and optical recordings. 5-HT is known to depress synaptic transmission at this synapse (Buchanan & Grillner, 1991). Using the intact lamprey spinal cord preparation, we show that 5-HT may depress synaptic transmission independently of [Ca2+]i regulation mechanisms.