Microdialysis coupled on-line to capillary liquid chromatography with tandem mass spectrometry for monitoring acetylcholine in vivo.
Capillary liquid chromatography-mass spectrometry (cLC-MS) was coupled on-line to microdialysis sampling to monitor endogenous acetylcholine (ACh) from the rodent brain. In vivo microdialysate sampled at 0.6 microL/min from the striatum of ketamine or chloral hydrate anesthetized rats was loaded onto a sample loop and then injected onto a approximately 5 cm long strong cation exchange (SCX) capillary column. A step gradient was used to separate the analyte from ionization suppressing salts contained in dialysate in 2.4 min. Sampling coupled on-line with cLC-MS allowed for high temporal resolution (data points at 2.4 min intervals), good reproducibility (10-15% relative standard deviation, R.S.D.), and sensitive limits of detection (0.04 nM or 8 amol injected). The method successfully monitored basal and stimulated levels (induced by increased K+ concentrations) of ACh from the anesthetized rat without necessitating perfusion of an acetylcholinesterase (AChE) inhibitor. Absolute and percent basal levels of ACh from rats receiving different anesthetics were also compared.
Shackman
Holly M
HM
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA.
Shou
Minshan
M
Cellar
Nicholas A
NA
Watson
Christopher J
CJ
Kennedy
Robert T
RT
D000109:Acetylcholine / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D000758:Anesthesia; D000778:Anesthetics, Dissociative; D018686:Anesthetics, Intravenous; D000818:Animals; D002138:Calibration; D053801:Capillary Electrochromatography; D002697:Chloral Hydrate; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D007202:Indicators and Reagents; D007649:Ketamine; D008297:Male; D013058:Mass Spectrometry; D017551:Microdialysis / Q000295:instrumentation* / Q000379:methods*; D009862:Online Systems; D051381:Rats; D017207:Rats, Sprague-Dawley; D012015:Reference Standards; D015203:Reproducibility of Results
D000778:Anesthetics, Dissociative; D018686:Anesthetics, Intravenous; D002800:Cholinesterase Inhibitors; D007202:Indicators and Reagents; D002697:Chloral Hydrate; D007649:Ketamine; D000109:Acetylcholine