Temporal neurotransmitter conditioning restores the functional activity of adult spinal cord neurons in long-term culture

Abstract

The ability to culture functional adult mammalian spinal-cord neurons represents an important step in the understanding and treatment of a spectrum of neurological disorders including spinal cord injury. Previously, the limited functional recovery of these cells, as characterized by a diminished ability to initiate action potentials and to exhibit repetitive firing patterns, has arisen as a major impediment to their physiological relevance. In this report we demonstrate that single temporal doses of the neurotransmitters serotonin, glutamate (N-acetyl-DL-glutamic acid) and acetylcholinechloride leads to the full electrophysiological functional recovery of adult mammalian spinal-cord neurons, when they are cultured under defined serum-free conditions. Approximately 60% of the neurons treated regained their electrophysiological signature, often firing single, double and, most importantly, multiple action potentials.

Preparation of Adult Spinal Cord Motor Neuron Cultures Under Serum-Free Conditions

“…This supports the view that mature SCMNs are strongly dependent on their interactions with neurotrophins, cAMP, and soluble factors from their target tissue. A similar trophic dependence of mature SCMNs has also been described by Das et al ( 27,28 ) . Adult rat SCMNs survive in the presence of a complex cocktail of neurotrophins ( 27 ) and they reach electrical functionality in the presence of neurotransmitters ( 28 ) .…”

“…However, other molecules may be involved in the survival of SCMNs. For instance, Das et al ( 27,28 ) showed that a more complex mixture of factors (BDNF, GDNF, neurotrophin-3, heparin sulfate, cardiotrophin-1, vitronectin, and acidic fi broblast growth factor) is necessary for the maintenance of adult SCMNs under their conditions.…”

“…In our experience, PDL gives the best results. Others have shown that the cell growthpromoting organosilane substrate, N-1 (3[trilnethoxysilyl] propyl)-diethylenetriainine, coated on a glass surface is necessary for the survival of these neurons ( 27,28 ) . PDL should not be used after one freeze-thaw cycle.…”

Regeneration and characterization of adult mouse hippocampal neurons in a defined in vitro system

“…A timed application of a limited number of growth factors to the culture medium substantially enhanced cell survival, as opposed to prolonged application. Our research shows that neurons cultured using this technique retain their phenotypic properties (Das et al, 2008). The use of synthetic substrates, such as DETA, allows for reproducible and precise quantification of the culture substrate properties.…”

Patterned cardiomyocytes on microelectrode arrays as a functional, high information content drug screening platform

“…Surface modification with SAMs is also compatible with advanced photolithography methods[30, 34]. Studies have also shown that cells survive on these surfaces for extended periods of time[35, 36], do not migrate off the patterned areas[34] and exhibit the typical morphology and physiology of the specific cell type[16, 37]. …”