Monitoring neurotransmitter release from isolated retinas using online microdialysis-capillary electrophoresis.
Release of neurotransmitters and other primary amine-containing analytes from intact, isolated larval salamander (Ambystoma tigrinum) retinas maintained in a 6.5-microL perfusion chamber was monitored using online microdialysis-capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). Primary amines were derivatized online with o-phthaldialdehyde (OPA) and beta-mercaptoethanol. With the use of overlapping injections, the perfusate was sampled every approximately 10 s. Although separation conditions were optimized using 20 mM hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD) for a number of important neuromessengers including D- and L-serine, D- and L-asparate, glutamate, GABA, serotonin, dopamine, norepinephrine, and taurine, only glutamate (0.48 +/- 0.27 microM), GABA (0.25 +/- 0.12 microM), taurine (5.5 +/- 2.1 microM), and l-serine (2.8 +/- 1.0 microM) were identified in the perfusate. Elevated levels of glutamate, GABA, and taurine were detected during stimulation with 60 mM K+. This method is the first to directly sample multiple neurotransmitters from perfused, isolated retinas and to observe changes in efflux of these neurotransmitters as a result of pharmacological stimulation.
O'Brien
Kylie B
KB
Department of Chemistry, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Esguerra
Manuel
M
Miller
Robert F
RF
Bowser
Michael T
MT
D000557:Ambystoma; D000818:Animals; D001692:Biological Transport; D019075:Electrophoresis, Capillary / Q000379:methods*; D007834:Lasers; D017551:Microdialysis / Q000379:methods*; D018377:Neurotransmitter Agents / Q000032:analysis* / Q000378:metabolism / Q000494:pharmacology; D009862:Online Systems; D010477:Perfusion; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D047408:gamma-Cyclodextrins / Q000737:chemistry
D018377:Neurotransmitter Agents; D047408:gamma-Cyclodextrins; C479659:hydroxypropyl-gamma-cyclodextrin