Applied electric fields (EFs) have previously been presented as a potential method of inducing functional recovery after neural trauma. To date, most of this research has focused on the application of a direct current (DC) stimulus to produce the desired EF and induce neuronal growth. We propose that high duty-cycle alternating current (AC) stimulation is capable of inducing similar EFs within the spinal cord and eliciting a neural response with the added benefits of increased field propagation and lower power consumption. Through ex vivo tissue testing of porcine spinal columns and Xenopus laevis cell cultures, 80% duty-cycle AC stimulation was compared to DC stimulation for efficacy in field generation and induction of neurite growth. Results from ex vivo measurement show that AC stimulation is capable of producing EFs of greater magnitudes over an increased distance in the spinal cord than DC stimulation at the same current magnitude. Furthermore, stimulation of Xenopus laevis neuronal cultures with 80% duty-cycle rectangular waves indicated a significant increase in neurite length as compared to non-stimulated controls and cathodal preference, growth that was statistically similar to DC-stimulated cells. These results suggest high duty-cycle stimulation modalities to be applicable and perhaps preferable to DC stimulation in electrically mediated neuronal therapies.

In this work we quantified the in vitro calibration relationships between high frequency electrical stimulation and GABA and glutamate release in both mature retinoic acid differentiated P19 neurons and immortalized embryonic cortical cells engineered to express glutamic acid decarboxylase, GAD65. Extracellular glutamate and GABA was quantified by 2D gas chromatography and time of flight mass spectrometry after stimulation at varying amplitudes and frequencies. Amplitude sweeps resulted in a linear calibration for P19 neurons; the level of neurotransmitter varied over one order of magnitude from ~ 200 pg/neuron to ~ 1.2 ng/neuron for glutamate and ~ 1 ng/neuron to ~ 2 ng/neuron for GABA, depending on the stimulation amplitude. Frequency sweeps resulted in a peak release at 250 Hz for glutamate and 400 Hz for GABA in P19 cells. The GABA transporter inhibitor, nipecotic acid, increased extracellular GABA levels and decrease glutamate. In contrast the embryonic cortical cells had a strongly nonlinear dependency of release on stimulation amplitude, and a weak dependence on frequency. These cells had roughly equal extracellular glutamate and GABA levels after stimulation despite the expression of GAD65. In addition glutamate and GABA levels were insensitive to nipecotic acid. These results demonstrate an ability to calibrate and tune neurotransmitter release from neural cells using high frequency stimulation parameters.

Microstimulation of neural tissue has become a widely-used technique for controlling neuronal responses with local electric fields as well as a therapeutic intervention for nervous system disorders such as epilepsy and Parkinson's disease. Of those afflicted by neurological diseases, many are or become tolerant to existing pharmaceuticals and are left with little recourse. Little is known about the necessary design criteria or efficacy of a hybrid neural prosthesis. Assessment of the potential clinical value of a hybrid electro-chemical neural prosthesis was performed through in vitro verification using a prototype microstimulator and P19 cell cultures. We constructed a printed circuit board (PCB) microstimulator as a prototype of a CMOS microstimulator ASIC that was subsequently fabricated in the IBM 7RF 0.18 microm process. Measured results for the prototype are described in this work. An output impedance of 237 kOmega, voltage compliance of 11.3 V, and a linear constant-current output up to +/-600 microA make this microstimulator system a viable option for an implantable hybrid neural prosthesis. Hybrid prostheses could uniquely affect neural modulation with linear glutamate release at physiological amplitudes and frequencies.