We have identified a serotonin-sensitive K+ channel with novel properties. The channel is active at the testing potential; its gating is moderately affected by membrane potential and is not dependent on the activity of intracellular calcium ions. Application of serotonin to the cell body or intracellular injection of cyclic AMP causes prolonged and complete closure of the channel, thereby reducing the effective number of active channels in the membrane. The closure of the channel can account for the increases in the duration of the action potential, Ca2+ influx, and transmitter release which underlie behavioural sensitization, a simple form of learning.

Serotonin exerts a long-lasting excitatory action on sensory neurons of Aplysia californica by decreasing outward K+ current. The depression of outward current delays repolarization of the action potential and extends the duration of Ca2+ influx into the presynaptic terminals, thereby contributing to the facilitation of transmitter release that underlies behavioral sensitization. We have extended the analysis of serotonin's action and find that it acts on a specific serotonin-sensitive K+ current (S current), which is different from the early K+ current ('A), the delayed K+ current ('K), the Ca2+-dependent K+ current (IC), and the muscarine-sensitive M current. The serotonin-sensitive current in these cells persists when IA and IK are reduced by conditioning depolarization or channel-blocking agents. The S current is not activated by intracellular injection ofCa2+, nor is it affected by substitution of Ba2+ for Ca2+, a treatment that reduces Ic.Moreover, intracellular injection of cyclic AMP exerts an effect indistinguishable from that of serotonin. This observation and the insensitivity ofthe current to Ba2+ distinguishes the S current from M current. S current is activated at the resting potential and does not inactivate with steady-state depolarization. It is active sufficiently early during an action potential to contribute to the repolarization of the action potential and therefore accounts for the physiological effects of serotonin.A number ofneurotransmitters produce long-lasting excitatory or inhibitory synaptic actions in neurons of both invertebrates and vertebrates (1, 2). Some of these slow synaptic potentials involve changes in membrane currents that are different from the usual fast synaptic actions. Whereas fast excitatory synaptic actions typically result from the opening ofcation channels that allow movement ofNa+ and K+, several slow excitatory synaptic actions result from a transmitter's closing channels that are more selective for K+ (3-5).In the marine snail Aplysia, serotonin or activation of the presynaptic facilitating neurons, thought to release serotonin, produces a slow excitatory synaptic action by reducing a K+ current in the sensory neurons of the gill-withdrawal reflex pathway (6, 7). This slow synaptic action is thought to be responsible for presynaptic facilitation of transmitter release, which underlies behavioral sensitization, a simple form of learning. The serotonin-modulated K+ current contributes importantly to repolarizing the action potential in the sensory neurons. Depression ofthis K+ current leads to a broadening ofthe action potential, which increases the influx ofCa2" and thereby enhances transmitter release from the terminals (6, 7). Several -lines of evidence indicate that serotonin or a related substance is the presynaptic facilitating transmitter and that it produces its action by a cyclic AMP-mediated phosphorylation of the K+ channel or of a protein associated with the channel (8-11).In this paper we have extended the analysis of the decreased K+ curre...

We have found that two endogenous neuropeptides in Aplysia, the small cardioactive peptides SCPA and SCPB, facilitate synaptic transmission from siphon mechanosensory neurons and enhance the defensive withdrawal reflex that these sensory neurons mediate. Single-channel recording revealed that these peptides close a specific K+ channel, the S channel, which is sensitive to cAMP. Moreover, the peptides increase cAMP levels in these sensory neurons. This reduction in K+ current slows the repolarization of the action potential in these cells, which increases transmitter release. In these actions, the SCPs resemble both noxious sensitizing stimuli, which enhance the reflex, and serotonin. Bioassay of HPLC fractions of abdominal ganglion extracts and immunocytochemistry indicate that both the SCPs and serotonin are present in the ganglion and are found in processes close to the siphon sensory neurons, suggesting that these transmitters may be involved in behavioral sensitization. Recent evidence suggests that one group of identified facilitatory interneurons, the L29 cells, does not appear to contain either the SCPs or serotonin but may use yet another facilitatory transmitter.Thus, it appears that several transmitters can converge to produce presynaptic facilitation in the sensory neurons of the defensive withdrawal reflex. AU of the transmitters studied here, the SCPs and serotonin, act via an identical molecular cascade: cAMP-dependent closure of the S-K' channel, broadening of the presynaptic action potential, and facilitation of transmitter release.Peptide transmitters have been a focus of interest in neurobiology because they often have modulatory effects on behavior (1). However, it has only rarely been possible to relate the behavioral action of a neuropeptide to its cellular action on specific nerve cells (2-4, 38). We here report that two molluscan neuropeptides increase the gain of the gill and siphon withdrawal reflex of Aplysia by presynaptically facilitating transmission from the sensory neurons of the reflex pathway. By combining single-channel and biochemical analyses, we have found that this facilitation involves the cAMP-mediated closure of a specific K+ channel, the recently described S channel (5).This defensive withdrawal reflex in Aplysia is enhanced during two simple forms of learning: sensitization and classical conditioning (6,7). In each case, a noxious stimulus produces presynaptic facilitation of synaptic transmission from the afferent neurons in the reflex, the mechanosensory neurons in the abdominal ganglion that innervate the siphon, to interneurons and motoneurons (8, 9). Although application of serotonin to the abdominal ganglion stimulates the facilitatory actions of noxious stimuli (10,11), there is now evidence from the immunocytochemical studies of Ono and McCaman (12) and Kistler et al. (13) that at least one set of facilitatory interneurons, the L29 cells (14), is not serotonergic. Thus transmitters other than serotonin are involved in sensitization. We were therefore ...

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Selected actions of neurotransmitters and hormones on ion channels in nerve and muscle cells are now thought to be mediated by cyclic AMP-dependent protein phosphorylation. Although the cyclic AMP-dependent protein kinase (cAMP-PK) affects the cellular properties of several neurones, its mode of action at the single-channel level has not been characterized. In addition, little is known about the identity or subcellular localization of the phosphoproteins that control channel activity and, in particular, whether the critical substrate proteins are cytoplasmic or membrane-associated. In Aplysia sensory neurones, serotonin produces a slow modulatory synaptic potential mediated by cAMP-PK that contributes to presynaptic facilitation and behavioural sensitization. Previously, we have found that serotonin acts on cell-attached membrane patches to produce prolonged all-or-none closures of a specific class of K+ channels (S channels) whose gating is weakly dependent on voltage and independent of intracellular calcium. We demonstrate here that in cell-free membrane patches from Aplysia sensory neurones, the purified catalytic subunit of cAMP-PK produces all-or-none closures of the S channel, simulating most (but not all) aspects of the action of serotonin on cell-attached patches. This result suggests that protein kinase acts on the internal surface of the membrane to phosphorylate either the channel itself or a membrane-associated protein that regulates channel activity.