Article

Defined Electrical Stimulation Emphasizing Excitability for the Development and Testing of Engineered Skeletal Muscle

Division of Neurobiology, Physiology and Behavior, University of California Davis, Davis, California, USA.
Tissue Engineering Part C Methods (Impact Factor: 4.64). 11/2011; 18(5):349-57. DOI: 10.1089/ten.TEC.2011.0364
Source: PubMed

ABSTRACT

Electrical stimulation is required for the maturation of skeletal muscle and as a way to nondestructively monitor muscle development. However, the wrong stimulation parameters can result in electrochemical damage that impairs muscle development/regeneration. The goal of the current study was to determine what aspect of an electrical impulse, specifically the pulse amplitude or pulse width, was detrimental to engineered muscle function and subsequently how engineered muscle responded to continuous electrical stimulation for 24 h. Acute stimulation at a pulse amplitude greater than six-times rheobase resulted in a 2.4-fold increase in the half-relaxation time (32.3±0.49 ms vs. 77.4±4.35 ms; p<0.05) and a 1.59-fold increase in fatigability (38.2%±3.61% vs. 60.6%±4.52%; p<0.05). No negative effects were observed when the pulse energy was increased by lengthening the pulse width, indicating electrochemical damage was due to electric fields at or above six-times rheobase. Continuous stimulation for 24 h at electric fields greater than 0.5 V/mm consistently resulted in ∼2.5-fold increase in force (0.30±0.04 kN/m² vs. 0.67±0.06 kN/m²; p<0.05). Forty per cent of this increase in force was dependent on the mammalian target of rapamycin (RAP) complex 1 (mTORC1), as RAP prevented this portion of the increase in force (CON=0.30±0.04 kN/m² to 0.67±0.06 kN/m² compared with RAP=0.21±0.01 kN/m² to 0.37±0.04 kN/m²; p<0.05). Since there was no increase in myosin heavy chain, the remaining increase in force over the 24 h of stimulation is likely due to cytoskeletal rearrangement. These data indicate that electrochemical damage occurs in muscle at a voltage field greater than six-times rheobase and therefore optimal muscle stimulation should be performed using lower electric fields (two- to four-times rheobase).

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Available from: Alastair Khodabukus, Mar 06, 2015
    • "However, the dimensions of these grooves were generally at the micron scale, which poorly approximates the diameter of myotubes in vivo (30–100 μm) [20] [21] [22]. Significantly, these 2D micropatterned substrates have been used only as tools for investigating mechanisms of cell function, and have not yet been developed into implantable tissue engineering constructs [23] [24]. Finally, 3D tubular scaffolds with an aligned pore structure and biologically relevant pore sizes appear to be promising for skeletal muscle tissue engineering, which generally requires a pore size of 100–150 μm; however, these dimensions are much larger than the cellular scale (∼10 μm) [18] [25]. "
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    • "In the future, the in vitro skeletal muscle tissues will have to be adapted and refined, for example, by co-culturing with motor neurons to improve the comparability of the tissue-engineered constructs with in vivo muscle. Recently, Khodabukus and Baar reported that continuous 24-h EPS to 14-day culture tissue constructs at pulse amplitude of 1 V/mm and width of 4 ms, which corresponded to 100%Pt, resulted in a 2-fold increase in force27, suggesting that high %Pt may be required to increase force production when using short-duration, acute EPS. However, we demonstrated here that low %Pt was effective for long-duration, chronic EPS, as we achieved a 4.5-fold increase in force by low %Pt EPS culture (Fig. 4a) without a reduction in force production due to electrochemical damage. "
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