Short-term synaptic plasticity compensates for variability in number of motor neurons at a neuromuscular junction

Abstract

We studied how similar postsynaptic responses are maintained in the face of interindividual variability in the number of presynaptic neurons. In the stomatogastric ganglion of the lobster, Homarus americanus, the pyloric (PY) neurons exist in variable numbers across animals. We show that each individual fiber of the stomach muscles innervated by PY neurons received synaptic input from all neurons present. We performed intracellular recordings of excitatory junction potentials (EJPs) in the muscle fibers to determine the consequences of differences in the number of motor neurons. Despite the variability in neuron number, the compound electrical response of muscle fibers to natural bursting input was similar across individuals. The similarity of total synaptic activation was not due to differences in the spiking activity of individual motor neurons across animals with different numbers of PY neurons. The amplitude of a unitary EJP in response to a single spike in a single motor neuron also did not depend on the number of PY neurons present. Consequently, the compound EJP in response to a single stimulus that activated all motor axons present was larger in individuals with more PY neurons. However, when axons were stimulated with trains of pulses mimicking bursting activity, EJPs facilitated more in individuals with fewer PY neurons. After a few stimuli, this resulted in depolarizations similar to the ones in individuals with more PY neurons. We interpret our findings as evidence that compensatory or homeostatic regulatory mechanisms can act on short-term synaptic dynamics instead of absolute synaptic strength.

Figure 1.

The nerve-muscle preparation of the stomatogastric system used in this study. A, Schematic of the STNS and muscle indicating intracellular and extracellular recording sites. B, Schematic of the lobster stomach showing the location of the PY-innervated muscles p2, p8, and p10 in the pylorus. C, Photomicrograph of the surgically isolated muscles with intact innervation. CoG, Commissural ganglion; OG, oesophageal ganglion; PD, pyloric dilator.

Figure 2.

Electrical muscle responses during spontaneous rhythmic pyloric activity. A, Simultaneous recordings of two different fibers in the p2 muscle, one from the anterior end, and one from the middle. B, Simultaneous recordings of a p2 and a p8 fiber. C, Simultaneous recordings of a p2 and a p10 fiber. Extracellular nerve recordings in all panels show the phasing of the pyloric rhythm. The magnified insets indicate the 1:1 relationship of EJPs in the paired recordings (dashed lines).

Figure 3.

Recruitment of presynaptic PY neuron axons in response to nerve stimulation with gradually increasing amplitude. A, Overlaid traces of EJP responses in p2. Gradually increasing stimulation amplitude led to stepwise increases in EJP amplitude, indicating successive recruitment of additional PY axons. B, Plot of EJP amplitude and stimulation voltage from the same experiment.

Figure 4.

Similarity of electrical responses of p2 fibers to spontaneous rhythmic pyloric input across preparations with different numbers of PY neurons. A, Intracellular p2 fiber recording, indicating the measurements taken. B, Box and scatter plots of the electrical response parameters indicated in A (3 PY neurons: n = 7; 4 PY neurons: n = 15; 5 PY neurons: n = 17). None of the parameters showed significant differences across preparations with different numbers of PY neurons.

Figure 5.

Similarity of PY neuron activity across preparations during spontaneous pyloric rhythms. A, Simultaneous recordings of a PY neuron soma in the STG, a p2 muscle fiber, and two motor nerves during spontaneous rhythmic pyloric activity. B, Box and scatter plots of PY burst parameters (3 PY neurons: n = 13; 4 PY neurons: n = 21; 5 PY neurons: n = 12).

Figure 6.

EJP parameters across preparations with different numbers of PY neurons. A, Examples of unitary EJPs (one PY axon activated) and compound EJPs (all PY axons activated) in response to single stimulation pulses (3 PY neurons: n = 9; 4 PY neurons: n = 15; 5 PY neurons: n = 17). B, Box and scatter plots of EJP amplitudes in unitary and compound responses across preparations with different numbers of PY neurons. C, Unitary EJP indicating the measurements taken to describe the time course. D, Box and scatter plots of unitary EJP time course measurements from the same data shown in B. *p < 0.05, **p < 0.01.

Figure 7.

Short-term synaptic dynamics in compound EJP responses. A, Examples of responses to repeated train stimulations of all PY axons. Shown are the 24th and 25th trains in preparations with different numbers of PY neurons. The dashed line indicates the peak voltage of the first response in the train. B, Box and scatter plots of compound EJP amplitudes for the first and last pulses in the last (25th) train across preparations with different numbers of PY neurons (3 PY neurons: n = 7; 4 PY neurons: n = 12; 5 PY neurons: n = 16).C, Box and scatter plots of the ratios between the compound EJP amplitudes in response to the last and first pulse of the last (25th) train. *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 8.

A, Overlaid compound EJP recordings from a stimulation protocol with a conditioning train and single test pulses at varying intervals, showing the synaptic enhancement and the time course of recovery. The dashed line indicates the EJP amplitude in response to the first pulse in the conditioning train. B, Plot of the mean response difference (n = 5) between test pulse and first pulse in the conditioning train for EJP peak voltages and preceding baseline voltages, as a function of test pulse interval. Note that summation ceases after a few hundred microseconds as the baseline voltage in between pulses returns to normal, while the increase of EJP amplitude persists for several seconds. C, Overlaid EJP recordings from paired-pulse stimulations with varying intervals (10 ms–2 s). Unitary and compound EJP recordings are from the same experiment. D, Mean paired-pulse response amplitude ratios as a function of interstimulus intervals (n = 5). Both unitary and compound responses show similar paired-pulse ratios at all interstimulus intervals.