Localization of D1 dopamine receptors on live cultured striatal neurons by quantitative fluorescence microscopy.
Single neurons in culture express a heterogeneity of neurotransmitter receptor subtypes. The study of the effects of neurotransmitters on neuronal function is complicated by this heterogeneity. It would therefore be useful to be able to identify live neurons that express the receptors of interest and then use these neurons for functional studies. We have used quantitative fluorescence microscopy to identify single live striatal neurons that express D1 dopamine receptors. The binding of the fluorescent D1 dopamine receptor antagonist bodipy-SCH 23390 was measured in 2-3-week-old primary striatal cultures derived from fetal rats (embryonic day 18). Binding of bodipy-SCH 23390 to live neurons was displaced by (+)-butaclamol, dopamine or SCH 23390, indicating that it specifically labelled D1 dopamine receptors. However, the fraction of bodipy-SCH 23390 binding that was specific varied substantially among individual neurons indicating heterogeneity of D1 dopamine receptor expression. Interestingly, bodipy-SCH 23390 also specifically labelled discrete spots of receptors on the neuronal processes. This technique should prove useful in the study of the effects of dopaminergic drugs on neuronal function in primary culture.
Hoyt
K R
KR
Department of Pharmacology, University of Pittsburgh, School of Medicine, PA 15261, USA.
Reynolds
I J
IJ
D000143:Acids / Q000032:analysis; D000818:Animals; D001552:Benzazepines / Q000378:metabolism; D001667:Binding, Competitive / Q000502:physiology; D001896:Boron Compounds; D002451:Cell Compartmentation / Q000502:physiology; D002478:Cells, Cultured / Q000737:chemistry; D018492:Dopamine Antagonists / Q000494:pharmacology; D005456:Fluorescent Dyes; D007700:Kinetics; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000737:chemistry / Q000378:metabolism; D017072:Neostriatum / Q000166:cytology*; D016501:Neurites / Q000737:chemistry; D009474:Neurons / Q000737:chemistry* / Q000166:cytology / Q000648:ultrastructure; D051381:Rats; D017447:Receptors, Dopamine D1 / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000378:metabolism
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D000143:Acids; D001552:Benzazepines; D001896:Boron Compounds; D018492:Dopamine Antagonists; D005456:Fluorescent Dyes; D017447:Receptors, Dopamine D1