Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Poster Abstracts

37-POS

Board 19

NMDA Receptors and Large-Conductance Calcium-Activated Potassium Channels in

Enkephalinergic Neurons of Mouse Spinal Superficial Dorsal Horn

Eiko Kato

, Yuuichi Hori.

Dokkyo Medical University, Tochigi, Japan.

Enkephalin (Enk)-containing neurons are distributed at a high density in the superficial dorsal

horn (SDH) of the spinal cord. In the present experiments, we analyzed whether Enk-containing

neurons in the SDH express the

-methyl-

-aspartate receptor (NMDAR) 2B (NR2B) subunit

and large-conductance calcium-activated potassium (BK) channels, and how the activity of Enk-

containing neurons is regulated by NMDAR and BK channels.

The experiments were performed on 5- to 8-week-old male ICR mice. The lumbar spinal cord

was dissected, and transverse slices were prepared. Tight-seal whole-cell recordings were

obtained from neurons in the SDH. After the recordings, the neurons were collected, and single-

cell real-time RT-PCR was performed to analyze the expression profile of genes in each SDH

neuron.

Puff application of L-glutamate evoked an inward current at a holding potential of -70 mV. Upon

depolarizing the holding potential to 0 mV, outward current of long duration appeared after

initial inward current. The NR2B-selective NMDAR antagonist ifenprodil reduced the outward

current. The outward current was also abolished by the selective BK channels antagonist

iberiotoxin. Single-cell real-time RT-PCR analysis revealed that single SDH neurons expressing

the preproenkephalin mRNA also expressed the BK channel α-subunit, NR1, and NR2B subunit

mRNAs. Additionally, the neurons generating the outward current showed a significant tendency

to express BK channels α-subunit mRNA, which is consistent with the pharmacological results

of a BK channels antagonist.

Our experiments suggest that combining real-time RT-PCR with whole-cell recordings provides

a useful tool to analyze the functions of genetically characterized central nervous neurons at the

single-cell level and that Ca

influx through NMDAR may activate BK channels and

hyperpolarize Enk-containing neurons in the SDH.