AVS 50th International Symposium
    Biomaterial Interfaces Tuesday Sessions
       Session BI-TuP

Paper BI-TuP20
In-vitro Electrophysiological Comparison of Embryonic Hippocampal Neurons Grown in 2D and 3D Environments

Tuesday, November 4, 2003, 5:30 pm, Room Hall A-C

Session: Poster Session
Presenter: T. Xu, Clemson University
Authors: T. Xu, Clemson University
P. Molnar, Clemson University
C.A. Gregory, Clemson University
M. Das, Clemson University
J.J. Hickman, Clemson University
T. Boland, Clemson University
Correspondent: Click to Email
To compare electrophysiological differences between neurons cultured in 2D and 3D environments, neurons dissociated form embryonic rat brain were seeded onto type-I collage thin coating, collagen gel surface, and were entrapped randomly into collagen gel, which were set up for imitating a 2D culture environment, a defined 3D environment and a random 3D environment for neuron culture respectively. Double-immunostaining for MAP-2, a neuronal cell body and dendritic marker, and anti-neurofilament antibody, an axonal marker, was used to identify neuron morphology. Hippocampal neuron polarities and outgrowth of neurites were evaluated by confocal microscope images. The gels were optimized for neuronal 3D cultures by varying concentrations, porosities, and glial cell densities. Surface properties of the gels will be characterized by AFM. Whole-cell patch clamp experiments were carried out to investigate electrophysiology of hippocampal neurons cultured in different conditions. Improvements of patch clamp technique for neuronal 3D culture over traditional 2D culture were applied. Membrane and synaptic properties of neurons in response to their different culture conditions were recorded and compared. Our preliminary results show that hippocampal neurons cultured in 2D and 3D environments exhibited similar passive membrane properties and sodium and potassium currents. Repetitive firings of action potential were found in neurons cultured in 3D environment over 14 days. Results will be presented on functional synapse formation for neurons cultured in 3D environment as measured by the combination of the patch clamp technique and the sharp microelectrode technique. The results of this study indicate that embryonic hippocampal neurons retain a clearly neuronal electrophysiological phenotype in a engineered in-vitro 3D culture condition, which is holding potential in applications ranging from neural tissue engineering to providing active neuronal networks for neuro-computing.