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Electrophysiological Characteristics of Enteric Neurons Isolated from the Immortomouse

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Abstract

Background

Recently, two enteric neuronal cell lines, one fetal and the other post-natal (IM-PEN), have been developed from the H-2Kb-tsA58 transgenic mouse (immortomouse). However, their electrophysiological properties are not known. The goal of this study was to determine the electrical excitability and ionic conductance of the immortalized postnatal enteric neuronal (IM-PEN) cell line.

Methods

Whole cell patch clamp studies, immunohistochemistry and RT-PCR were performed on differentiated IM-PEN cells following propagation at 33 °C and differentiation at 37 °C.

Results

Differentiated IM-PEN cells stained positively for the neuron specific markers βIII-tubulin and PGP9.5. The mRNA for several ion channels expressed in enteric neurons were detected by RT-PCR. In current clamp, the resting membrane potential was −24.6 ± 2.1 mV (n = 6) for IM-FEN and −29.8 ± 0.9 mV (n = 30) for IM-PEN. Current injections from Vh −80 mV resulted in passive responses but not action potentials. Depolarizing pulses in the whole cell voltage clamp configuration from Vh −80 mV elicited small nifedipine-sensitive inward currents. Additionally, outward currents with slow deactivating tail currents were blocked by niflumic acid and low chloride solution. A volume-regulated anion current was elicited by hypo-osmotic solution and inhibited by 10 μM DCPIB. Growth with rabbit gastrointestinal smooth muscle did not yield significant differences in the active properties of the IM-PEN cell line. Transient expression of L-type Ca2+ channels produced large inward currents demonstrating a working mechanism for protein folding and transport.

Conclusion

The electrophysiological characteristics of IM-PEN cells suggest that chloride channels in IM-PEN cells play an important role in their resting state, and membrane trafficking of some of the ion channels may preclude their electrical excitability.

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Abbreviations

IM-FEN:

Immortalized fetal enteric neurons

IM-PEN:

Immortalized post-natal enteric neurons

DCPIB:

4-[(2-Butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]butanoic acid

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Acknowledgments

This work was supported by NIH grants DK46367, DA024009 and T32DA007027 and NIH-RO1 (DK080684, SS), VA-MERIT award (SS).

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Authors and Affiliations

  1. Department of Pharmacology and Toxicology, Medical College of Virginia Campus, Virginia Commonwealth University, 1112 E, Clay Street, McGuire Hall 317, Richmond, VA, 23298, USA

    Edward G. Hawkins, William L. Dewey & Hamid I. Akbarali

  2. Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, VA, USA

    John R. Grider

  3. Division of Digestive Diseases, Emory University, Atlanta, GA, USA

    Mallappa Anitha & Shanthi Srinivasan

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  1. Edward G. Hawkins
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  2. William L. Dewey
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  3. Mallappa Anitha
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  6. Hamid I. Akbarali
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Correspondence to Hamid I. Akbarali.

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Hawkins, E.G., Dewey, W.L., Anitha, M. et al. Electrophysiological Characteristics of Enteric Neurons Isolated from the Immortomouse. Dig Dis Sci 58, 1516–1527 (2013). https://doi.org/10.1007/s10620-013-2557-5

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  • DOI: https://doi.org/10.1007/s10620-013-2557-5

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