Differential role of inhibition in habituation of two independent afferent pathways to a common motor output

Abstract

Many studies of the neural mechanisms of learning have focused on habituation, a simple form of learning in which a response decrements with repeated stimulation. In the siphon-elicited siphon withdrawal reflex (S-SWR) of the marine mollusk Aplysia, the prevailing view is that homosynaptic depression of primary sensory afferents underlies short-term habituation. Here we examined whether this mechanism is also utilized in habituation of the tail-elicited siphon withdrawal reflex (T-SWR), which is triggered by an independent, polysynaptic afferent pathway that converges onto the same siphon motor neurons (MNs). By using semi-intact preparations in which tail and/or siphon input to siphon MNs could be measured, we found that repeated tail stimuli administered in the presence of a reversible conduction block of the nerves downstream of the tail sensory neurons (SNs) completely abolished the induction of habituation. Subsequent retraining revealed no evidence of savings, indicating that the tail SNs and their immediate interneuronal targets are not the locus of plasticity underlying T-SWR habituation. The networks closely associated with the siphon MNs are modulated by cholinergic inhibition. We next examined the effects of network disinhibition on S-SWR and T-SWR habituation using an Ach receptor antagonist d-tubocurarine. We found that the resulting network disinhibition disrupted T-SWR, but not S-SWR, habituation. Indeed, repeated tail stimulation in the presence of d-tubocurarine resulted in an initial enhancement in responding. Lastly, we tested whether habituation of T-SWR generalized to S-SWR and found that it did not. Collectively, these data indicate that (1) unlike S-SWR, habituation of T-SWR does not involve homosynaptic depression of SNs; and (2) the sensitivity of T-SWR habituation to network disinhibition is consistent with an interneuronal plasticity mechanism that is unique to the T-SWR circuit, since it does not alter S-SWR.

Figure 1.

T-SWR habituation does not occur when training occurs in the presence of a conduction block of the P-ACs. (A) Illustration of experimental protocol. (B) Intracellular recordings from siphon MNs in two separate preparations during T-SWR habituation training conducted in ASW (top) and while conduction via the P-ACs was blocked using a MgCl₂ conduction block (bottom). In ASW, the tail tap-evoked MN response showed substantial decrement after 10 taps, and this response recovered to baseline within 10 min. Following training under P-AC conduction block, the MN response showed no decrement. (C) Summary data from 10 experiments showing that repeated tail stimulation decrements the tail tap-evoked response in siphon MNs (HABITUATION) and that the same training conducted in the presence of a conduction block of the P-ACs (MgCl₂ + HAB) yields responses similar to nonhabituated controls (ASW).

Figure 2.

Lack of savings following habituation training in the presence of a conduction block of the P-ACs. (A) Illustration of experimental protocol. The letters a, b, and c signify the initial pre-tests, the pre-tests for the second phase of training, and the habituation training, respectively. (B) Ratio of post-test response following T-SWR habituation training in the presence of MgCl₂ conduction block or equivalent time point in ASW controls (b in above protocol) to initial pre-tests of tail tap-evoked MN responses (a in above protocol). Neither habituation training with the conduction block (solid bar; N = 6) nor time alone (open bar; N = 5) changed tap-evoked MN response. (C) Summary of 11 experiments showing overlapping habituation and recovery curves generated by naive preps (ASW) and those previously trained under conduction block (Pre-HAB).

Figure 3.

Network disinhibition enhances baseline T-SWR and S-SWR. (A) Illustration of experimental protocol. (B) Histograms depicting the enhancement of baseline T-SWR (left, N = 5) and S-SWR (right, N = 5) following 100 μM d-TC exposure. ASW treatment alone resulted in no change in baseline T-SWR (N = 5) or S-SWR (N = 5).

Figure 4.

Network disinhibition disrupts habituation of T-SWR. (A) Intracellular recording from siphon MNs during T-SWR habituation experiments under conditions of ASW control (A1) and d-TC disinhibition (A2). (A1) In ASW, the tail tap-evoked MN responses did not change across pre-tests. The response diminished across habituation trials. (A2) The tap-evoked MN response increased after addition of 100 μM d-TC to the bath. In addition, the response did not diminish across habituation trials. In this case, d-TC resulted in a marked increase in the tap-evoked response during the first three habituation trials. (B) Summary of 14 experiments showing the disruption of T-SWR habituation by d-TC disinhibition.

Figure 5.

Network disinhibition does not affect habituation of S-SWR. (A) Intracellular recording from siphon MNs during S-SWR habituation experiments under conditions of ASW control (A1) and d-TC disinhibition (A2). (A1) In ASW, the siphon tap-evoked MN responses did not change across pre-tests. The response diminished across habituation trials. (A2) The tap-evoked MN response increased after addition of 100 μM d-TC to the bath. However, the response similarly diminished across habituation trials. (B) Summary of 10 experiments showing no effect of d-TC disinhibition on habituation of S-SWR.

Figure 6.

Habituation of T-SWR does not generalize to S-SWR. (A) Illustration of experimental protocol. (B) Intracellular recordings from a single siphon MN in a preparation with which MN responses to both siphon (top) and tail (bottom) could be recorded. The MN response to siphon tap was unaltered by habituation of the T-SWR. (C) Summary data from five experiments showing that habituation of the T-SWR does not affect the S-SWR.

Figure 7.

Model circuits depicting possible mechanisms of habituation in the T-SWR and S-SWR. Hypothesized neural circuits underlying T-SWR (top) and S-SWR (bottom). The S-SWR circuit consists of both monosynaptic and polysynaptic components. Activity-dependent, homosynaptic depression occurs at synapses marked with asterisks (^*). Sites of blockade in our experiments are indicated by dashed lines.