Date of Award

1979

Degree Type

Thesis

Degree Name

Master of Science in Pharmacology and Toxicology

Specialization

Pharmacology

Department

Pharmacology and Toxicology

First Advisor

Stephen Nelson

Abstract

Mouse brain choline-0-acetyltransferase (E.C. 2.3.1.6) (ChAT) activity was studied in vitro using a lOOmM sodium phosphate buffer (pH 7.4) wash of a crude vesicular fraction containing solubilized ChAT and a washed crude vesicular fraction containing membrane bound ChAT. Both the solubilized and membrane bound forms of ChAT can acetylate choline linearly for 30 minutes. High concentrations of acetylcholine (Ach) can inhibit solubilized ChAT to a greater degree than the membrane bound enzyme form. FoFtY percent of the Ach synthesized by membrane bound ChAT survives hydrolysis in the presence of an excess of acetylcholinesterase (AchE), whereas none of the Ach synthesized by the solubilized enzyme form does. 4-(l-naphthylvinyl) pyridine inhibits solubilized ChAT more than the membrane bound form, either in vivo or in vitro, and reduces mouse locomotor activity by 80-90%. for 3 hours. Solubilized ChAT is totally sodium dependent whereas the membrane bound form is only partially sodium dependent. Membrane bound ChAT has ·both a high (Km 3.2 uM) and a low affinity (Km=0.48 mM) Michaelis constants as a function of added choline when velocity values are not corrected for acetylation of endogenous choline. Upon correction, only a low affinity Michaelis constant (Km=0.47 mM) is obtained for the membrane bound enzyme form, similar to that of solubilized ChAT (Km=.17 mM). The choline analogues triethylcholine and homocholine are acetylated preferentially by membrane bound ChAT. The results suggest the existence of a membrane bound form of ChAT in mouse brain which is able to synthesize Ach.

Download

Share

COinS
 
 

To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.