The use of exogenous electrical stimulation to promote nerve regeneration has achieved only limited success. Conditions impeding optimized outgrowth may arise from inadequate stimulus presentation due to differences in injury geometry or signal attenuation. Implantation of an electrically-conductive biomaterial may mitigate this attenuation and provide a more reproducible signal. In this study, a conductive nanofiller (single-walled carbon nanotubes [SWCNT]) was selected as one possible material to manipulate the bulk electrical properties of a collagen type I-10% Matrigel™ composite hydrogel. Neurite outgrowth within hydrogels (SWCNT or nanofiller-free controls) was characterized to determine if: 1) nanofillers influence neurite extension and 2) electrical stimulation of the nanofiller composite hydrogel enhances neurite outgrowth. Increased SWCNT loading (10–100-μg/ml) resulted in greater bulk conductivity (up to 1.7-fold) with no significant changes to elastic modulus. Neurite outgrowth increased 3.3-fold in 20-μg/mL SWCNT loaded biomaterials relative to the nanofiller-free control. Electrical stimulation promoted greater outgrowth (2.9-fold) within SWCNT-free control. The concurrent presentation of electrical stimulation and SWCNT-loaded biomaterials resulted in a 7.0-fold increase in outgrowth relative to the unstimulated, nanofiller-free controls. Local glia residing within the DRG likely contribute, in part, to the observed increases in outgrowth; but it is unknown which specific nanofiller properties influence neurite extension. Characterization of neuronal behavior in model systems, such as those described here, will aid the rational development of biomaterials as well as the appropriate delivery of electrical stimuli to support nerve repair.

Background γ-Aminobutyric acid type A (GABAA) receptors mediate important effects of intravenous general anesthetics. Photolabel derivatives of etomidate, propofol, barbiturates, and a neurosteroid get incorporated in GABAA receptor transmembrane helices M1 and M3 adjacent to intersubunit pockets. However, photolabels have not been consistently targeted at heteromeric αβγ receptors and do not form adducts with all contact residues. Complementary approaches may further define anesthetic sites in typical GABAA receptors. Methods Two mutation-based strategies, substituted tryptophan sensitivity and substituted cysteine modification–protection, combined with voltage-clamp electrophysiology in Xenopus oocytes, were used to evaluate interactions between four intravenous anesthetics and six amino acids in M1 helices of α1, β3, and γ2L GABAA receptor subunits: two photolabeled residues, α1M236 and β3M227, and their homologs. Results Tryptophan substitutions at α1M236 and positional homologs β3L231 and γ2L246 all caused spontaneous channel gating and reduced γ-aminobutyric acid EC50. Substituted cysteine modification experiments indicated etomidate protection at α1L232C and α1M236C, R-5-allyl-1-methyl-5-(m-trifluoromethyl-diazirinylphenyl) barbituric acid protection at β3M227C and β3L231C, and propofol protection at α1M236C and β3M227C. No alphaxalone protection was evident at the residues the authors explored, and none of the tested anesthetics protected γ2I242C or γ2L246C. Conclusions All five intersubunit transmembrane pockets of GABAA receptors display similar allosteric linkage to ion channel gating. Substituted cysteine modification and protection results were fully concordant with anesthetic photolabeling at α1M236 and β3M227 and revealed overlapping noncongruent sites for etomidate and propofol in β+–α– interfaces and R-5-allyl-1-methyl-5-(m-trifluoromethyl-diazirinylphenyl) barbituric acid and propofol in α+–β– and γ+–β– interfaces. The authors’ results identify the α+–γ– transmembrane interface as a potentially unique orphan modulator site.

Magnetic electrospun fibers are of interest for minimally invasive biomaterial applications that also strive to provide cell guidance. Magnetic electrospun fibers can be injected and then magnetically positioned in situ, and the aligned fiber scaffolds provide consistent topographical guidance to cells. In this study, magnetically responsive aligned poly-L-lactic acid electrospun fiber scaffolds were developed and tested for neural applications. Incorporating oleic acid-coated iron oxide nanoparticles significantly increased neurite outgrowth, reduced the fiber alignment, and increased the surface nanotopography of the electrospun fibers. After verifying neuron viability on two-dimensional scaffolds, the system was tested as an injectable three-dimensional scaffold. Small conduits of aligned magnetic fibers were easily injected in a collagen or fibrinogen hydrogel solution and repositioned using an external magnetic field. The aligned magnetic fibers provided internal directional guidance to neurites within a three-dimensional collagen or fibrin model hydrogel, supplemented with Matrigel. Neurites growing from dorsal root ganglion explants extended 1.4–3× farther on the aligned fibers compared with neurites extending in the hydrogel alone. Overall, these results show that magnetic electrospun fiber scaffolds can be injected and manipulated with a magnetic field in situ to provide directional guidance to neurons inside an injectable hydrogel. Most importantly, this injectable guidance system increased both neurite alignment and neurite length within the hydrogel scaffold.

Affecting approximately 17,000 new people each year, spinal cord injury (SCI) is a devastating injury that leads to permanent paraplegia or tetraplegia. Current pharmacological approaches are limited in their ability to ameliorate this injury pathophysiology, as many are not delivered locally, for a sustained duration, or at the correct injury time point. With this review, we aim to communicate the importance of combinatorial biomaterial and pharmacological approaches that target certain aspects of the dynamically changing pathophysiology of SCI. After reviewing the pathophysiology timeline, we present experimental biomaterial approaches to provide local sustained doses of drug. In this review, we present studies using a variety of biomaterials, including hydrogels, particles, and fibers/conduits for drug delivery. Subsequently, we discuss how each may be manipulated to optimize drug release during a specific time frame following SCI. Developing polymer biomaterials that can effectively release drug to target specific aspects of SCI pathophysiology will result in more efficacious approaches leading to better regeneration and recovery following SCI.

Three aligned, electrospun fiber scaffolds with unique surface features were created from poly-L-lactic acid (PLLA). Fibers without surface nanotopography (smooth fibers), fibers with surface divots (shallow pits), and fibers with surface pits (deeper pits) were fabricated, and fiber alignment, diameter, and density were characterized using scanning electron microscopy (SEM). Whole dorsal root ganglia (DRG) were isolated from rats and placed onto uncoated fibers or fibers coated with laminin. On uncoated fibers, neurite outgrowth was restricted by fibers displaying divoted or pitted nanotopography when compared to neurite outgrowth on smooth fibers. However, neurites extending from whole DRG cultured on laminin-coated fibers were not restricted by divoted or pitted surface nanotopography. Thus, neurites extending on laminin-coated fibers were able to extend long neurites even in the presence of surface divots or pits. To further explore this result, individual neurons isolated from dissociated DRG were seeded onto laminin-coated smooth, pitted, or divoted fibers. Interestingly, neurons on pitted or divoted fibers exhibited a 1.5-fold increase in total neurite length, and a 2.3 or 2.7-fold increase in neurite branching compared to neurons on smooth fibers, respectively. Based on these findings, we conclude that fiber roughness in the form of pits or divots can promote extension and branching of long neurites along aligned electrospun fibers in the presence of an extracellular matrix protein coating. Thus, aligned, electrospun fibers can be crafted to not only direct the extension of axons but to induce unique branching morphologies.