Localized gene transfer into organotypic hippocampal slice cultures and acute hippocampal slices.
Viral vectors derived from herpes simplex virus, type-1 (HSV), can transfer and express genes into fully differentiated, post-mitotic neurons. These vectors also transduce cells effectively in organotypic hippocampal slice cultures. Nanoliter quantities of a virus stock of HSVlac, an HSV vector that directs expression of E. coli beta-galactosidase (beta-gal), were microapplied into stratum pyramidale or stratum granulosum of slice cultures. Twenty-four hours later, a cluster of transduced cells expressing beta-gal was observed at the microapplication site. Gene transfer by microapplication was both effective and rapid. The titer of the HSVlac stocks was determined on NIH3T3 cells. Eighty-three percent of the beta-gal forming units successfully transduced beta-gal after microapplication to slice cultures. beta-Gal expression was detected as rapidly as 4 h after transduction into cultures of fibroblasts or hippocampal slices. The rapid expression of beta-gal by HSVlac allowed efficient transduction of acute hippocampal slices. Many genes have been transduced and expressed using HSV vectors; therefore, this microapplication method can be applied to many neurobiological questions.
Casaccia-Bonnefil
P
P
Program of Anatomy and Cell Biology, State University of New York-Health Science Center at Brooklyn 11203.
Benedikz
E
E
Shen
H
H
Stelzer
A
A
Edelstein
D
D
Geschwind
M
M
Brownlee
M
M
Federoff
H J
HJ
Bergold
P J
PJ
D000818:Animals; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000502:physiology; D018259:Herpesvirus 1, Human / Q000201:enzymology / Q000235:genetics; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism*; D066298:In Vitro Techniques; D009474:Neurons / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014161:Transduction, Genetic; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
D001616:beta-Galactosidase