Effects of pertussis toxin and G alpha-protein-specific antibodies on phosphoinositide hydrolysis in rat brain membranes after cholinergic denervation and hippocampal sympathetic ingrowth
K. Kolasa et al., Effects of pertussis toxin and G alpha-protein-specific antibodies on phosphoinositide hydrolysis in rat brain membranes after cholinergic denervation and hippocampal sympathetic ingrowth, EXP NEUROL, 161(2), 2000, pp. 724-732
Cholinergic denervation of the hippocampal formation, via medial septal les
ions, induces peripheral noradrenergic fibers, originating from the superio
r cervical ganglion, to grow into the hippocampus. We have previously repor
ted that cholinergic denervation and hippocampal sympathetic ingrowth diffe
rentially affect guanosine-5'-O-(3-thiotriphosphate)- as well as guanosine-
5'-O-(3-thiotriphosphate) + carbachol-stimulated polyphosphoinositide hydro
lysis, suggesting an alteration in G proteins and/or the entire receptor co
mplex, To examine the type of G protein which may be involved in these effe
cts, rat dorsal hippocampal membranes were preincubated with pertussis toxi
n in the presence of guanosine-5'-O-(3 -thiotriphosphate) and guanosine-5'-
O-(3-thiotriphosphate) + carbachol. Pertussis toxin reduced guanosine-5'-O-
(3-thiotriphosphate) in all groups, while guanosine-5'-O-(3-thiotriphosphat
e) + carbachol-stimulated phosphoinositide hydrolysis was reduced in contro
ls and animals without sympathetic ingrowth but not in animals with hippoca
mpal sympathetic ingrowth. This suggests that pertussis toxin-sensitive G:p
roteins may be involved in the mediation of phosphoinositide hydrolysis. To
confirm this hypothesis, membranes were preincubated with antibodies to G
alpha o and Gq/11. The Go antibody significantly decreased guanosine-5'-O-(
3-thiotriphosphate) in all groups, while guanosine-5'-O-(3-thiotriphosphate
) + carbachol-stimulated phosphoinositide hydrolysis was reduced only in hi
ppocampal sympathetic ingrowth, Impairment of guanosine-5'-O-(3-thiotriphos
phate) and carbachol-stimulated phosphoinositide hydrolysis was also decrea
sed in all groups when preincubated with Gq/11 antibody. To determine wheth
er hippocampal sympathetic ingrowth or cholinergic denervation altered the
concentration of various G proteins, immunoblotting methodology was utilize
d. Gq/11 concentrations were found to be equivalent among groups. The densi
ty of Gel, Go2, and Go3 isoforms was significantly increased in the choline
rgic denervation, while in the hippocampal sympathetic ingrowth only group
Go3 was significantly increased. When assessed as total Go protein, density
was increased significantly only in the cholinergic denervation group. Ove
rall, these results suggest that hippocampal sympathetic ingrowth and choli
nergic denervation induce alterations in phosphoinositide hydrolysis throug
h both the Gq/11 and the Go proteins and that the coupling between muscarin
ic receptor and G protein is the possible site which affects changes in pho
sphoinositide turnover. Our results also suggest that cholinergic denervati
on and hippocampal sympathetic ingrowth may mediate phosphoinositide hydrol
ysis through an effect on different isoforms of the Same G protein. (C) 200
0 Academic Press.