Intraspinal microstimulation and diaphragm activation after cervical spinal cord injury

Abstract

Intraspinal microstimulation (ISMS) using implanted electrodes can evoke locomotor movements after spinal cord injury (SCI) but has not been explored in the context of respiratory motor output. An advantage over epidural and direct muscle stimulation is the potential of ISMS to selectively stimulate components of the spinal respiratory network. The present study tested the hypothesis that medullary respiratory activity could be used to trigger midcervical ISMS and diaphragm motor unit activation in rats with cervical SCI. Studies were conducted after acute (hours) and subacute (5-21 days) C₂ hemisection (C2Hx) injury in adult rats. Inspiratory bursting in the genioglossus (tongue) muscle was used to trigger a 250-ms train stimulus (100 Hz, 100-200 μA) to the ventral C₄ spinal cord, targeting the phrenic motor nucleus. After both acute and subacute injury, genioglossus EMG activity effectively triggered ISMS and activated diaphragm motor units during the inspiratory phase. The ISMS paradigm also evoked short-term potentiation of spontaneous inspiratory activity in the previously paralyzed hemidiaphragm (i.e., bursting persisting beyond the stimulus period) in ∼70% of the C2Hx animals. We conclude that medullary inspiratory output can be used to trigger cervical ISMS and diaphragm activity after SCI. Further refinement of this method may enable "closed-loop-like" ISMS approaches to sustain ventilation after severe SCI.NEW & NOTEWORTHY We examined the feasibility of using intraspinal microstimulation (ISMS) of the cervical spinal cord to evoke diaphragm activity ipsilateral to acute and subacute hemisection of the upper cervical spinal cord of the rat. This proof-of-concept study demonstrated the efficacy of diaphragm activation, using an upper airway respiratory EMG signal to trigger ISMS at the level of the ipsilesional phrenic nucleus during acute and advanced postinjury intervals.

Keywords: diaphragm function; hypoglossal respiratory activity; phrenic motor nucleus; rat; respiration.

Fig. 1.

Cervical ISMS activates the ipsilateral diaphragm A: schematic diagram illustrates the EMG recording sites relative to the placement of the stimulating electrode (S). The stimulating electrode was placed in the immediate vicinity of the phrenic motor nucleus, and ISMS was initiated via a trigger signal based on inspiratory tongue (genioglossus) EMG activity. LI, left intercostal. B: pre-C2Hx representative EMG recordings from the genioglossus, extensor carpi radialis (ECR), and both sides of the diaphragm showing baseline (prestimulation) activity and a period of genioglossus-triggered cervical ISMS (100 Hz, 200 μA; 0.3-ms pulse duration, 250-ms train duration) represented by a gray box in the ∫tongue trace. C: expanded trace of the EMG recording from the left diaphragm during the period of ISMS shows each stimulus artifact, indicated by asterisks, and each subsequent MUAP. D: overlay of each elicited MUAP, aligned by the stimulus artifact, demonstrates constant latency and amplitude.

Fig. 2.

Examples of raw EMG activity recorded in the tongue (genioglossus), forelimb (ECR), and the left and right hemidiaphragm during ISMS. A: representative examples before C2Hx. B: stimulus-triggered averages of EMG activity show prominent MUAPs in the forelimb and ipsilateral diaphragm. C: stimulus-triggered averages of EMG activity after neuromuscular blockade. In B and C, the stimulus artifact is highlighted by the gray boxes. Stimulus-triggered averages were scaled to the same values and represent 25 stimulus triggers. In these examples, ISMS was delivered at 100 Hz and 200 μA during expiration.

Fig. 3.

ISMS after acute C2Hx. A: illustration of the electrode placement relative to the C2Hx lesion. The solid dark bar represents the injury site; the dashed lines represent severed pathways. B: examples of EMG activity in the right and left diaphragm before C2Hx. C: examples of left and right diaphragm EMG activity after acute C2Hx. Note that activity is abolished in the left diaphragm. In B and C, 3 spontaneous breaths are shown followed by 3 breaths during triggered cervical ISMS (solid bars; 100 Hz, 200 μA). D: stimulus-triggered averages of right and left diaphragm EMG activity obtained from the period of ISMS highlighted by dashed boxes in B and C. Stimulus-triggered averages were scaled to the same values and represent 25 stimulus triggers. In D, the stimulus artifact is highlighted by the gray boxes.

Fig. 4.

Impact of ISMS on tracheal pressure. A: representative examples of tracheal pressure and left diaphragm EMG activity before and during cervical ISMS (100 Hz, 200 μA). The period of ISMS is indicated by the solid bars. Note the negative deflection in tracheal pressure during ISMS. B: overlay plot further illustrates the tracheal pressure before stimulation (solid line) and during ISMS (dashed line). C: average change in tracheal pressure during lung inflation at prestimulation baseline and during ISMS. Data are shown for the spinal-intact condition (left) and after acute C2Hx (right). *P < 0.05, **P < 0.005.

Fig. 5.

ISMS after subacute C2Hx. A: representative diaphragm EMG activity after subacute C2Hx. In the example traces, genioglossus-triggered ISMS was delivered during the breaths marked by the black bars. B: histological section of the C₂ spinal cord stained with cresyl violet. The example demonstrates an anatomically complete hemilesion extending to the midline of cervical cord. C: stimulus-triggered averages from the ipsilesional (solid line) and contralesional diaphragm (dashed line); data were obtained from the period indicated by the dashed boxes in A. These traces illustrate activation of the diaphragm ipsilateral to the C2Hx lesion. D: average change in tracheal pressure during lung inflation at prestimulation baseline and during ISMS. *P < 0.05.

Fig. 6.

Example recordings illustrate short-term potentiation of ipsilateral diaphragm EMG activity following ISMS. A: tongue and ipsilateral (left) diaphragm EMG activity after acute C2Hx, during ISMS, and immediately after cessation of ISMS. Note that clear phasic (inspiratory) activity can be seen after ISMS, whereas the baseline showed no such activity. B: a similar response can be observed in a subacute C2Hx animal. In these examples, the ISMS was triggered by the inspiratory genioglossus EMG signal and was delivered at 100 Hz for 1 min of respiratory efforts.