Synaptic Facilitation and Behavioral Dishabituation in Aplysia: Dependence on Release of Ca2+ from Postsynaptic Intracellular Stores, Postsynaptic Exocytosis, and Modulation of Postsynaptic AMPA Receptor Efficacy

Abstract

Sensitization and dishabituation of the defensive withdrawal reflex in Aplysia have been ascribed to presynaptic mechanisms, particularly presynaptic facilitation of transmission at sensorimotor synapses in the CNS of Aplysia. Here, we show that facilitation of sensorimotor synapses in cell culture during and after serotonin (5-HT) exposure depends on a rise in postsynaptic intracellular Ca 2ϩ and release of Ca 2ϩ from postsynaptic stores. We also provide support for the idea that postsynaptic AMPA receptor insertion mediates a component of synaptic facilitation by showing that facilitation after 5-HT offset is blocked by injecting botulinum toxin, an exocytotic inhibitor, into motor neurons before application of 5-HT. Using a reduced preparation, we extend our results to synaptic facilitation in the abdominal ganglion. We show that tail nerve shock-induced facilitation of siphon sensorimotor synapses also depends on elevated postsynaptic Ca 2ϩ and release of Ca 2ϩ from postsynaptic stores and recruits a late phase of facilitation that involves selective enhancement of the AMPA receptor-mediated synaptic response. To examine the potential role of postsynaptic exocytosis of AMPA receptors in learning in Aplysia, we test the effect of injecting botulinum toxin into siphon motor neurons on dishabituation of the siphon-withdrawal reflex. We find that postsynaptic injections of the toxin block dishabituation resulting from tail shock. Our results indicate that postsynaptic mechanisms, particularly Ca 2ϩ -dependent modulation of AMPA receptor trafficking, play a critical role in synaptic facilitation as well as in dishabituation and sensitization in Aplysia.

Whereas short-term facilitation is presynaptic, intermediate-term facilitation involves both presynaptic and postsynaptic protein kinases and protein synthesis

“…Previous studies have found that short-term facilitation by 1-min 5-HT involves covalent modifications but does not require protein synthesis (Montarolo et al 1986;Martin et al 1997). By contrast, facilitation by 10-min 5-HT involves both covalent modifications and protein synthesis (Li et al 2005;Villareal et al 2007), suggesting that it represents a form of intermediate-term facilitation. However, the relative contributions of pre-and postsynaptic mechanisms have not been fully explored with either protocol.…”

“…For example, how are the postsynaptic mechanisms recruited, and what signaling molecules are involved? One possibility is that the postsynaptic mechanisms are recruited in parallel with the presynaptic mechanisms by activation of pre-and postsynaptic 5HT receptors (Li et al 2005;Fulton et al 2008;Villareal et al 2007Villareal et al , 2009. Alternatively, recent studies have found that activation of presynaptic 5-HT receptors enhances spontaneous release of glutamate, which then activates metabotropic glutamate receptors to recruit the postsynaptic mechanisms of intermediate-term facilitation (Jin et al 2007(Jin et al , 2008 and initiate a cascade that may culminate in synaptic growth during long-term facilitation (Jin et al 2010).…”

“…On the one hand, under some circumstances short-and long-term plasticity can be produced independently, suggesting that they may be induced in parallel (Emptage and Carew 1993). On the other hand, in both Aplysia and hippocampus an intermediate-term stage of plasticity has been identified that usually involves protein but not RNA synthesis and structural alterations but not synaptic growth, and therefore might form a bridge between short-and long-term plasticity (Ghirardi et al 1995;Winder et al 1998;Sutton and Carew 2000;Sutton et al 2001;Kim et al 2003;Li et al 2005Li et al , 2009Villareal et al 2007). That idea in turn suggests that aspects of the different stages of plasticity may be induced in series, similar to the states in artificial "cascade" models of memory storage (Fusi et al 2005).…”

Spontaneous transmitter release recruits postsynaptic mechanisms of long-term and intermediate-term facilitation in Aplysia

“…4B). However, DNQX significantly reduced facilitation after washout of the 5HT (F[1,112] = 5.29, P < 0.05 compared with control), suggesting that expression of the later part of facilitation may involve up-regulation of AMPA-like receptors (13). To examine this mechanism more directly, we expressed in the motor neuron the Aplysia homolog of the AMPA receptor subunit GluR1, ApGluR1 (GenBank accession no.…”

“…In addition, we found that this signaling is engaged quite early, during an intermediate-term stage that incorporates elements of both shortand long-term plasticity (11,12), and therefore might form part of a bridge or cascade connecting them. The intermediate stage is also the first stage to involve both presynaptic molecular mechanisms and postsynaptic mechanisms including IP3, Ca 2+ , protein synthesis, and membrane insertion of AMPA-like receptors (1,2,13,14). Postsynaptic Ca 2+ and protein synthesis are in turn necessary for long-term facilitation, in part through retrograde signaling (15).…”