Glutamate and aspartate measurements in individual planaria by rapid capillary electrophoresis.

INTRODUCTION

Planaria present a unique model organism for studying primitive central nervous systems. The major mammalian excitatory neurotransmitters, glutamate and aspartate, have previously been measured in planaria via high pressure liquid chromatography (HPLC). A faster extraction and analysis procedure using capillary electrophoresis (CE) was developed which confirms the presence of these amino acids in single planaria homogenates.

METHOD

Following homogenization and centrifugation of individual planaria in hydrochloric acid/acetonitrile, glutamate and aspartate were derivatized with naphthalene-2, 3-dicarboxaldehyde (NDA). The labeled amino acids were measured using capillary electrophoresis with laser-induced fluorescence (CE-LIF).

RESULTS

CE-LIF electropherograms were generated in less than 1 min. The mean ± S.D. amounts of glutamate and aspartate were 1200 ± 500 and 1900 ± 700 pmol/mg-planarian (n=22), respectively. Spiked average recoveries of glutamate and aspartate were 96% and 91%, respectively.

DISCUSSION

The high-throughput method provides the ability to quantitate changes in excitatory neurotransmitters under developmental or stimulatory conditions. The capability to monitor multiple neurotransmitter levels offers the opportunity to correlate behavioral responses with biochemical changes in planaria.