electrically stimulated muscle cells to pace contraction by using a 6-well plate 31. In a circular-shaped area, a uniform EF cannot be intuitively created by two electric potentials due to different electrical resistances originated from the length difference between the diameter of the circle and the length of any parallel chord of the bottom circular chamber where cells are cultured. Even though larger cell yields have been recently achieved by scaling up the rectangular shaped microchamber with increased cell culture area 29, 30, a large fraction of the circular shaped TCPS dish is still unutilised. As a result, a large portion of the cell culture area on the dish is unused, leading to low cell yield and poor cell product recovery. Further, a simple rectangular shaped cell culture microchamber is usually placed on a circular shaped tissue-culture polystyrene (TCPS) petri dish to generate the uniform EF. The small cross-section of the chamber limits the applicable electrical current and reduces the Joule heating that could be harmful to the cells.ĭespite the success of using microfluidic chips for electrical stimulation in recent studies, these microfluidic chips often require special fabrication procedures on cell culture dishes days prior to the actual experiment, limiting the adaptivity with common laboratory settings. Thus conventional electrotaxis studies usually employ a confined microfluidic chip in which cells are cultured in the bottom of the culture chamber 25, 26, 27, 28, 29, 30. The EF created through direct electrode stimulation is not uniform and cells are often exposed to toxic electrolysis products. Even though an electrical cue can direct cell migration comparable to that of chemical cues 22 and synergistically promote directional migration with other physical factors such as shear stresses 23, electrotaxis is less well studied than chemotaxis, possibly due to the lack of experimental tools for convenient EF stimulation comparable to a boyden chamber (transwell chamber) that is routinely used for chemotaxis 24.Ĭonventional in vitro electrical stimulations were commonly performed either by direct stimulation using electrodes, or stimulation in a microfluidic chamber with salt bridges. Gaining a better understanding of signalling pathways demands a reliable and convenient electrical stimulation platform for microscopy imaging and cell product recovery with subsequent biochemical analysis. Further investigations are required to clarify the functional roles of EF sensory proteins and signalling networks in regulating the electrotaxis phenomena. Various membrane receptors 6, 7, 8, 9, 10 or ion channels 11, 12, 13, 14, 15 have been suggested to act as EF sensors and initiate many intracellular signalling cascades in different cell types 8, 13, 16, 17, 18, 19, 20, 21. Numerous cellular signalling pathways have been regulated under electric field (EF) stimulation. The electrotaxis and dcEF stimulation have played pivotal roles in physiological processes such as embryonic development, neurogenesis, morphogenesis, and wound healing 1, 2, 3, 4, 5. Cells demonstrate directional migration (electrotaxis) or orientation-change (electro-alignment) in response to a physiological dcEF in both in vitro and in vivo settings. The CAD based inserts can be easily scaled up (i.e., 100 mm dishes) to further increase effective stimulation area percentages, and also be implemented in commercially available cultureware for a wide variety of EF-related research such as EF-cell interaction and tissue regeneration studies.Ī weak direct-current electric field (dcEF) exists at the tissue level due to the transepithelial potential difference established by the tissue polarity 1. In particular, NIH/3T3 mouse embryonic fibroblast cells are used to validate the performance of the 3D designed Poly(methyl methacrylate) (PMMA) inserts in a circular-shaped 6-well plate. A uniform EF with a coefficient of variation (CV) of 1.2% in the 6-well plate can be generated with an effective stimulation area percentage of 69.5%. To address this challenge, we develop a three-dimensional (3D) computer-aided designed (CAD) polymeric insert to create uniform EF in circular shaped multi-well culture plates. However, in a circular-shaped device, it is difficult to create uniform EF from two electric potentials due to different electrical resistances originated from the length difference between the diameter of the circle and the length of any parallel chord of the bottom circular chamber where cells are cultured. Applying uniform electric field (EF) in vitro in the physiological range has been achieved in rectangular shaped microchannels.
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