Multidirectional effect of low-intensity neuromuscular electrical stimulation on gene expression and phenotype in thigh and calf muscles after one week of disuse

Low-intensity neuromuscular electrical stimulation (NMES) is often used as an alternative to exercise and high-intensity electrical stimulation to prevent the loss of muscle mass, strength, and endurance in spaceflight and in patients with severe chronic diseases. This study investigated the effects of a one-week disuse, both with and without low-intensity neuromuscular electrical stimulation – a safe (non-traumatic) approach to prevent the loss of muscle mass, on the functional capacities and gene expression in thigh and calf muscles. This study assessed the efficiency of low-intensity (~10% of maximal voluntary contraction) electrical stimulation in preventing the negative effects of 7-day disuse (dry immersion without [see a related dataset GSE271607] and with daily stimulation) on the strength and aerobic performance of the ankle plantar flexors and knee extensors, mitochondrial function in permeabilized muscle fibers, and the proteomic (quantitative mass spectrometry-based analysis) and transcriptomic (RNA-sequencing) profiles of the soleus muscle and vastus lateralis muscle. Application of electrical stimulation during dry immersion prevented a decrease in the maximal strength and a slight reduction in aerobic performance of the knee extensors, as well as a decrease in maximal (intrinsic) ADP-stimulated mitochondrial respiration and changes in the expression of genes encoding mitochondrial, extracellular matrix, and membrane proteins in the vastus lateralis muscle. In contrast, for the ankle plantar flexors/soleus muscle, electrical stimulation had a positive effect only on maximal mitochondrial respiration, but slightly accelerated the decline in the maximal strength and muscle fiber cross-sectional area, which appears to be related to the activation of inflammatory genes. The data obtained open up broad prospects for the use of low-intensity electrical stimulation to prevent the negative effects of disuse for “mixed” muscles, meanwhile, the optimization of the stimulation protocol is required for “slow” muscles. Ten males participated in a 7-day dry immersion with NMES (DI+NMES, ages 26–39 years; height 1.74 m [1.70–1.76 m]; body mass 73 kg [70–79 kg]; and body mass index 24 kg/m² [23–26 kg/m²]). Motor neuron firing rate plays an important role in the activation of calcium-dependent signaling and the expression of muscle-specific genes. Therefore, in our study, both low (25 Hz) and high (50 Hz) stimulation frequencies were used during DI+NMES (in the first and second NMES daily session, respectively). Every day at 11:00 (7 sessions in total), low-intensity low-frequency electrical stimulation was applied to the calf and thigh muscles for 40–45 min (bipolar symmetrical rectangular pulses [1 ms, 25 Hz] for 1 s, followed by a 2-s pause). Every day at 17:00 (7 sessions in total), low-intensity high-frequency electrical stimulation was applied to the calf and thigh muscles for 10 min (carrier frequency of 2-kHz bipolar symmetrical pulses [10 ms separated by 10-ms pause] with bursts of 50 Hz for 10 s, followed by a 50-s pause). The burst time during high-frequency NMES was longer than that during low-frequency NMES (10 s vs. 1 s). Therefore, to increase the pain tolerance during high-frequency NMES, the carrier frequency (2-kHz) was used. Muscle samples were taken using a Bergstrom needle with aspiration under local anesthesia (2 ml of 2% lidocaine) from the medial part of the vastus lateralis (VL) and soleus (Sol) muscles of the left leg 14 days before and 6 days after the start of disuse (DI and DI+NMES) at 10:00, 3 hours after a standardized very light breakfast (5.2 g protein, 2.7 g fat, 55 g carbohydrates, 1253 kJ). Strand-specific RNA libraries were prepared using the NEBNext Ultra II RNA kit (New England Biolabs, USA) and single-end sequencing was done by a NextSeq 550 analyzer (Illumina, USA).