Panel A shows schematic of an ex vivo spinal cord hindlimb preparation with proprioceptor, motor neuron, neuromuscular junction, and nerve stimulation pathways. Panel B shows electrophysiological traces of tibialis anterior and gastrocnemius EMG responses during common peroneal nerve stimulation in WT preparations. Panel C shows electrophysiological traces of tibialis anterior and gastrocnemius EMG responses during common peroneal stimulation in C3 deficient and CD47 deficient preparations. Panel D shows bar graphs quantifying H responses and H response amplitudes during common peroneal nerve stimulation. Panel E shows scatter plots comparing M response and H response latencies between genotypes following common peroneal nerve stimulation. Panel F shows bar graphs quantifying H responses and H response amplitudes during tibial nerve stimulation. Panel G shows scatter plots comparing M response and H response latencies between genotypes following tibial nerve stimulation.
Inappropriate proprioceptive sensory synapses on motor neurons induce responses in antagonistic muscles. (A) Schematic of the ex vivo spinal cord–hindlimb preparation. Suction electrodes were placed on the CP and Tb nerves to perform en passant stimulation. Bipolar concentric needle electrodes recorded EMG activity from the TA and Gs muscles. A motor neuron and a neuromuscular junction (NMJ) are shown in green. A proprioceptor with its muscle spindle and the Ia central synapse on motor neurons are shown in purple. (B) Simultaneous EMG recordings from TA and Gs muscles following CP nerve stimulation in a WT spinal cord under control aCSF solution and after exposure to NBQX (20 μM) and D-AP5 (50 μM). Red traces show the average of five trials (grey). Black arrowhead denotes a stimulus artifact. (C) Traces from C3−/− (left) and CD47−/− (right) mice in control aCSF. Green arrows point to homonymous muscle H-reflex, while blue arrows point to inappropriate H-responses from the antagonistic Gs muscle following CP nerve stimulation. (D) Amplitude of H-reflex induced by CP nerve stimulation in the homonymous TA muscle (left; CP nerve → TA EMG) and the H-response from the antagonistic Gs muscle (right; CP nerve → Gs EMG). (E) Latency measurements of M-responses evoked by CP nerve stimulation in the TA muscle (grey circles), as well as H-reflex in the TA muscle following CP nerve stimulation (red squares), and antagonistic H-responses in the Gs muscle following CP nerve stimulation (red circles). (F) Amplitude of H-reflex induced by Tb nerve stimulation in the homonymous Gs muscle (left; Tb nerve → Gs EMG) and the H-response from the antagonistic TA muscle (right; Tb nerve → TA EMG). Each data point corresponds to a single animal (WT: N = 5; C3−/−: N = 4; CD47−/−: N = 3 mice). Significance: *P < 0.05, **P < 0.01, ***P < 0.001; one-way ANOVA with multiple comparisons using Bonferroni’s test. ns: no significance. (G) Latency measurements of M-responses evoked by Tb nerve stimulation in the Gs muscle (grey circles), as well as H-reflex in the Gs muscle following Tb nerve stimulation (red squares), and antagonistic H-responses in the TA muscle following Tb nerve stimulation (red circles). One-way ANOVA with multiple comparisons using Bonferroni’s test; ns: no significance.