ReviewNeural responses to electrical stimulation in 2D and 3D in vitro environments
Introduction
According to the U.S Department of Health and Human Services over 20 million people suffer or have suffered from peripheral nerve damage (Services 2014). This damage compromises the physical integrity of the Peripheral Nervous System (PNS), impacting the bi-directional transmission of electrical signals from limbs and organs to the Central Nervous System (CNS). Patients with peripheral nerve injuries (PNIs) present symptoms that involve numbness, loss of sensation, and even mechanical dysfunction resulting in lost productivity and increased socioeconomic isolation and burden. Minor and non-localized injuries with low risk of functional loss (e.g. nerve damage caused by diabetes) are generally not treated, relying on the patient’s self-regeneration capacities for recovery. On the other hand, severe nerve damage, e.g. traumatic injuries that compromise the nerve continuity, often require clinical interventions that represent around 200,000 repairs annually (Kehoe et al., 2012). Unfortunately, in these cases only 50% of patients regain useful function and few will completely recover resulting in long term disabilities, thus motivating the need for new and rationally defined treatment options.
Further, in the CNS approximately 1.7 million U.S. citizens suffer from traumatic brain injuries (TBIs) with an economic burden of up to $76.5 billion (Faul et al., 2010; Coronado et al., 2010, 2012; Cancelliere et al., 2017). In addition to TBI, there are 12,000–20,000 new spinal cord injuries (SCIs) per year (Raspa et al., 2016; Mazzoleni et al., 2013; Sahni and Kessler, 2010; Jazayeri et al., 2015). With the inclusion of neurodegenerative disorders, such as Parkinson’s and Alzheimer’s disease that also have limited therapeutic interventions, the impact of neural dysfunction on the quality of life for millions of people remains limited. In the CNS, regenerative capacity is limited by inhibitory barriers, such as glial scar formation and immune cell-mediated inflammation, motivating new therapeutic approaches such as electrical stimulation (ES) to modulate cellular function for improved restoration of function (Grinsell and Keating, 2014).
The application of ES for biological applications has been examined for over two hundred years, with increasing interest as technology and methods for cell culture and interfacing have expanded (Bresadola, 1998; Galvani, 1791). It is known that endogenous electrical cues drive development and morphogenesis, cellular communication networks, and neuronal and glial cellular signaling (Baer and Colello, 2016; Funk, 2015; Adams and Levin, 2013; Cao et al., 2013; Nuccitelli, 2003; Hotary and Robinson, 1992; Stump and Robinson, 1983); thus harnessing native machinery to modulate cellular activity in a controlled manner is attractive. ES has shown promise in modulating neural cell morphology (Shen et al., 2016; Yan et al., 2014; Huang et al., 2012b; Koppes et al., 2011; Lee et al., 2009) and impacting neural stem cell (NSC) fate (George et al., 2017; Hassarati et al., 2016; Cao et al., 2013; Jahanshahi et al., 2013; Park et al., 2011). However, a main limitation in successful translation to the clinic is the varied parameters, materials, and heterogeneous cell populations that limit a defined and mechanistic understanding of how to best modulate neural and support cells for specific therapeutic advantages.
This review focuses on the impact of ES in vitro, in 2D and 3D microenvironments to modulate neural outgrowth and function, support cell (e.g. glia), and NSC behavior. A consideration for how ES is applied, which materials are utilized for cellular and culture media interfacing, as well as limitations in mechanistic understanding is discussed. A mechanistic understanding will enable more targeted approaches in ES for specific cellular responses within heterogenous tissues to be realized. A brief outlook on alternative stimulation regimes, such as mechanical and light stimulation is also introduced. For some further reading on in vivo stimulation, readers are directed elsewhere (Koppes et al., 2014a; Ren et al., 2016; Wenger et al., 2016; Young, 2015; Angeli et al., 2014; Beuter et al., 2014; Kalia et al., 2013; Miocinovic et al., 2013; Hardenacke et al., 2012; Deuschl et al., 2011).
Section snippets
Historical overview
The link between electricity and biology began with the work of Luigi Galvani in the 18th century (Bresadola, 1998). By eliciting muscle contractions via ES of the sciatic nerve in a frog post mortem, Galvani was the first to demonstrate the excitability of nervous tissue. The influence of an applied electric field (EF) has been well documented for polarized cell populations including neurons, muscle, and glia, but all cells may be influenced by charge and ionic gradients (Batista Napotnik et
Neurons
Neurons synapse with other neurons to form neural networks, muscles to control movement, or convert sensory input intro electrochemical signals. A fundamental property of neurons is their ability to plastically respond to endogenous and exogenous stimuli (Sasmita et al., 2018; Lu et al., 2019; Werner and Stevens, 2015). Recent advances identified a plethora of neuronal subpopulations characterized by regional and epigenetic differences granting fine-tuned control over function (Lake et al., 2016
Future directions and other modes of stimulation
In addition to direct electrical coupling, a number of strategies for indirect coupling are being implemented to apply stimulation to in vitro cultures. These types of mechanisms will enable targeting of specific areas or spatiotemporal control over specific subpopulations of cells, enabling a minimization of side effects, off site targeting, or electrode byproducts at the site of application. An electromagnetic field can be induced via two parallel plates in close proximity (0.5–2 mm) to
Conclusions
Over the past few decades, the application of exogenous ES to modulate neural cell behavior for regenerative and therapeutic applications has gained momentum. Bioelectrical communication between neurons and support cell populations for restoration of function and wound healing has great potential for aiding recovery and regeneration, however there are still hurdles that require attention for the clinical relevance to be realized. As described above, the cell type and heterogeneity, species and
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