Muscle plasticity in rat following spinal transection and chronic intraspinal microstimulation

Abstract

Intraspinal microstimulation (ISMS) employs electrical stimulation of the ventral grey matter to reactivate paralyzed skeletal muscle. This work evaluated the transformations in the quadriceps muscle that occurred following complete transection and chronic stimulation with ISMS or a standard nerve cuff (NCS). Stimulation was applied for 30 days, 4 h/day. Both methods induced significant increases in time-to-peak tension (ISMS 35%, NCS 25%) and half rise-time (ISMS 39%, NCS 25%) compared to intact controls (IC). Corresponding increases in type-IIA myosin heavy chain (MHC) and decreases in type-IID MHC were noted compared to IC. These results were unexpected because ISMS recruits motor units in a near-normal physiological order while NCS recruits motor units in a reversed order. Spinal cord transection and 30 days of stimulation did not alter either recruitment profile. The slope of the force recruitment curves obtained through ISMS following transection and 30 days of stimulation was similar to that obtained in intact animals, and 3.4-fold shallower than that obtained through NCS. The transformations observed in the current work are best explained by the near maximal level of motor unit recruitment, the total daily time of activity and the tonic nature of the stimulation paradigm.

Figure 1. Summary of twitch width measurements

A) TTP lengthened in both NCS and ISMS legs compared to NCC and ISC control legs as well as IC legs. B) 1/2RT lengthened in both NCS and ISMS legs compared to NCC, ISC and IC legs. C) 1/2FT lengthened in NCS and ISMS legs as compared to the NCC and ISMS legs. Only the ISMS legs showed significant lengthening of 1/2FT when compared to the IC leg. (a) different from intact control, (b) different from contralateral control.

Figure 2. Myosin heavy chain (MHC) labelling with immunohistochemistry

Representative examples of MHC staining from rectus femoris muscles at 5X magnification. A) Intact control legs show a mixed phenotype with a preponderance of MHC type-IIB and unstained -IID fibres. B, D) Following spinal cord transection, unstimulated nerve cuff control (B) and intraspinal microwire control (D) legs show increases in MHC type-IIB fibre content and decreases in MHC type-IIA fibre content. C, E) Thirty days of stimulation through a nerve cuff (C) or intraspinal microwires (E) causes an increase in MHC type-IIA fibre content and a decrease in MHC type-IID content. A mean of 432.3 ± 96.1 rectus femoris muscle fibres were analyzed in each leg for a total of 12970 fibres.

Figure 3. Summary of MHC immunohistochemistry results from the deep portion of the muscle

Stimulation with either NCS or ISMS caused a fast-to-slow conversion as demonstrated by increases in type-IIA fibre content and corresponding decreases in type-IID fibre content as compared to the IC leg. Control NCC and ISC legs showed a slow-to-fast conversion as demonstrated by increases in type-IIB fibre content and decreases in type-IIA fibre content. (a) different from intact control, (b) different from contralateral control

Figure 4. Whole muscle analysis of the MHC IIA content

Whole muscle images immunolabeled for MHC IIA (A) were binarized so that stained pixels were rendered completely black and all other pixels completely white (B). Image analysis software was used to quantify black pixels and express them as a fraction of all pixels within the muscle boundary. (C) The area of the muscle containing MHC IIA increased in stimulated ISMS and NCS legs compared to control legs ISC and NCC and to the IC leg. (a) different from intact control, (b) different from contralateral control.

Figure 5. Pattern of force recruitment after chronic ISMS and NCS

Normalized force recruitment curves obtained through A) 6 nerve cuff electrodes and B) 5 ISMS microwires, after 30 days of stimulation. Graphs show force recruited by one second long 50 pps trains of randomly assigned amplitudes. The mean slope of the force recruited from 10% to 90% of peak force was 3.4 fold sharper in the nerve cuff group. Maximal force evoked was 6.29 N ± 4.02 N through the nerve cuff and 1.64 N ± 0.39 N through the ISMS microwires.