Long-term sensitization training produces spike narrowing in Aplysia sensory neurons

Abstract

Both short- and long-term sensitization of withdrawal reflexes of Aplysia are attributable at least in part to facilitation of the sensorimotor synapse. Previously, short-term synaptic facilitation has been associated with spike broadening and no change in temporal dynamics of burst transmission. In the present study, we examined whether long-term sensitization (LTS) is also associated with spike broadening and whether long-term synaptic facilitation is accompanied by changes in temporal dynamics. The results indicate that the temporal dynamics of the sensorimotor synapse are preserved after long-term facilitation. However, in contrast to short-term sensitization, LTS was accompanied by spike narrowing. The spike narrowing was observed both in centrally triggered spikes in isolated ganglia and in peripherally triggered spikes in reduced tail preparations. In addition, in reduced tail preparations, fewer spike failures in the afferent discharge of sensory neurons occurred in response to tail stimulation after ipsilateral LTS. Collectively, the results reveal that long-term sensitization affects the spike waveform of sensory neurons and enhances the sensory neuron responses to peripheral stimuli, but does not modify the synaptic dynamics of homosynaptic depression.

Figure 1.

Protocol for sensitization training. A, All training stimuli were delivered on a randomly chosen side of the lateral body wall. Testing stimuli were delivered on the ipsilateral and contralateral sides of the tail, at an area different from the one where training stimuli were delivered. B, Four-day training for LTS started immediately after a session of baseline behavioral testing. Twenty-four hours after the end of training, an identical behavioral testing session was used to assess LTS on the ipsilateral side of trained animals versus the contralateral side of trained animals or the untrained animals.

Figure 2.

Long-term sensitization training induces parallel effects on behavior of animals and efficacy of sensorimotor synapses. A, Average data on the changes in duration of siphon-withdrawal reflex 24 h after 4 d LTS training. The tail-elicited siphon withdrawal reflex is sensitized only when the ipsilateral side of the tail is tested. When the contralateral side is tested, the reflex is similar to that elicited by stimulation of untrained animals, indicating that long-term memory for sensitization is unilateral. B, Average data on the peak amplitude of monosynaptic EPSPs recorded from the cell body of motor neurons after triggering spikes in presynaptic sensory neurons. In pleural-pedal ganglia of ipsilaterally trained animals, EPSPs had significantly larger amplitude than in synapses from contralaterally trained and untrained animals. Inset, example traces of EPSPs. **p < 0.01.

Figure 3.

Long-term sensitization does not modify the temporal dynamics of sensorimotor synapses. A1, Example traces of the relative depression of motor neuron responses during 1 s, 10 Hz stimulation of spikes at presynaptic sensory neurons. These recordings come from animals that had received ipsilateral or contralateral LTS training, or from untrained animals. Traces have been scaled to match in the peak amplitude of the first EPSP. A2, Average data of synaptic depression induced by 10 Hz stimulation. For each synapse tested, EPSP amplitudes were normalized to the amplitude of the first EPSP. There was no difference among the three groups on the depression achieved with 10 stimuli at 10 Hz. B1, Example traces of the relative depression of motor neuron responses during repeated stimulation at 10 s intervals. The traces (scaled as in A1) come from ipsilaterally trained, contralaterally trained, or untrained animals. B2, Average data of synaptic depression induced by 0.1 Hz stimulation. For each synapse tested, EPSP amplitudes were normalized to the amplitude of the first EPSP. There was no difference among the three groups on the depression achieved with 10 stimuli at 0.1 Hz.

Figure 4.

Long-term sensitization is associated with spike narrowing in ipsilateral sensory neurons. A1, In isolated ganglia preparations, one sharp electrode was used to trigger single spikes and another was used to record the spikes at the cell body of sensory neurons. A2, Example traces of spikes recorded from sensory neurons of pleural ganglia in ipsilaterally trained, contralaterally trained, and untrained animals. These traces are representative of average spike duration. A3, Average data from the three groups of preparations. Spike width was significantly reduced in ipsilateral sensory neurons compared with contralateral and untrained sensory neurons. B1, The reduced tail preparation includes the isolated tail and parapodia of the animal, with the left P9 nerve intact, connected to the left pleural-pedal ganglia. AC stimuli, similar to the ones used to assess the intact animal's behavior, are delivered to the skin of the tail through implanted silver-wire electrodes. The peripherally evoked spike activity is recorded passively from the cell body of responsive tail sensory neurons. B2, Example traces of spikes recorded from tail sensory neurons of ipsilaterally trained, contralaterally trained, and untrained animals. These traces are representative of average spike duration. Spikes were narrower in preparations from ipsilaterally trained animals. B3, Average data from the three groups of preparations. Spike width was significantly reduced in ipsilaterally trained animals compared with contralaterally trained and untrained animals. The difference in absolute spike widths between panels A3 and B3 is caused by the ∼7° difference in temperature. *p < 0.05.

Figure 5.

Long-term sensitization enhances the sensory neuron responses to cutaneous stimuli. A1, Example traces of sensory neuron spike activity in an ipsilaterally trained, a contralaterally trained, and an untrained animal in response to a 20 ms AC stimulus (i.e., 1.2 cycles at 60 Hz). A2, In 60% of ipsilaterally trained preparations, the 20 ms AC triggered spike doublets, versus only 14 and 17% of contralaterally trained and untrained preparations, respectively. The complementary percentages correspond to neurons that fired single spikes. B1, Example traces of sensory neuron spike activity in an ipsilateral, a contralateral, and an untrained preparation, in response to a 200 ms AC stimulus (i.e., 12 cycles at 60 Hz). A sensory neuron response included a spike failure if there were <12 spikes in the time-locked phase of the response. B2, In 100% of preparations from ipsilaterally trained animals, sensory neurons fired at least 12 spikes (i.e., no failures) (mean number ± SEM, 12.60 ± 0.25). In contrast, in 50% of preparations from contralateral and untrained animals, sensory neuron responses had at least one spike failure (contra, 10.50 ± 1.29; untrained, 9.50 ± 1.20). B3, Spike width was a significant predictor of the number of spikes sensory neurons would fire in response to a 200 ms peripheral stimulus. The line of best fit had a slope of −2.92.