Single-cell RT-PCR detects shifts in mRNA expression profiles of basal forebrain neurons during aging.
The medial septum and nucleus of the diagonal band (MS/nDB) contain cholinergic and GABAergic neuronal populations that have been identified based on immunohistochemical staining and/or electrophysiological properties. We explored the molecular diversity of MS/nDB neurons using single-cell reverse transcription-polymerase chain reaction (scRT-PCR) to assess gene expression profiles during aging in individual neurons acutely isolated from young (2-4 months) and aged (26-27 months) F344 rats. Neuronal gene expression profiles were characterized by detection of mRNAs for choline acetyltransferase (ChAT, cholinergic) and glutamate decarboxylase (GAD67, GABAergic), as well as mRNAs for calcium binding proteins (CaBPs) calbindin-D28k, calretinin and parvalbumin. Four major neuronal populations were identified: ChAT-positive (ChAT+) cells, GAD-positive (GAD+) cells, ChAT+/GAD+ cells and ChAT negative/GAD negative (ChAT-/GAD-) cells. With age, the percentage of cells expressing ChAT mRNA decreased from 53% in young to 40%, and the expression of GAD67 mRNA was reduced from 56 to 35% of the cells tested. The percentage of cells with detectable levels of both ChAT and GAD67 mRNA was reduced from 24% in young to 9% in aged. Concomitantly, the percentage of ChAT-/GAD- cells increased from 15 to 34% with age. Of the CaBPs, calretinin expression was observed most frequently in this study, and its detection decreased from 33 to 22% of the cells with age. Observations concerning the CaBPs were confirmed using in situ hybridization. These results suggest a shift in the mRNA expression profiles of MS/nDB neuronal populations during aging and exemplify the molecular diversity of cholinergic and GABAergic cells.
Han
Sun-Ho
SH
Department of Medical Pharmacology and Toxicology, College of Medicine, Texas A&M University System Health Science Center, College Station, TX 77843-1114, USA.
McCool
Brian A
BA
Murchison
David
D
Nahm
Sang-Soep
SS
Parrish
Alan R
AR
Griffith
William H
WH
D000375:Aging / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D064092:Calbindin 1; D064032:Calbindin 2; D064026:Calbindins; D002118:Calcium / Q000378:metabolism; D002795:Choline O-Acetyltransferase / Q000096:biosynthesis / Q000235:genetics; D002799:Cholinergic Fibers / Q000378:metabolism; D004790:Enzyme Induction; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D005968:Glutamate Decarboxylase / Q000096:biosynthesis / Q000235:genetics; D017403:In Situ Hybridization; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D008832:Microchemistry; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics*; D009473:Neuronal Plasticity; D009474:Neurons / Q000378:metabolism*; D010320:Parvalbumins / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D011916:Rats, Inbred F344; D020133:Reverse Transcriptase Polymerase Chain Reaction; D064030:S100 Calcium Binding Protein G / Q000096:biosynthesis / Q000235:genetics; D013687:Telencephalon / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000502:physiology
C576500:Calb1 protein, rat; C576512:Calb2 protein, rat; D064092:Calbindin 1; D064032:Calbindin 2; D064026:Calbindins; D007527:Isoenzymes; D009419:Nerve Tissue Proteins; D010320:Parvalbumins; D012333:RNA, Messenger; D064030:S100 Calcium Binding Protein G; D005680:gamma-Aminobutyric Acid; D002795:Choline O-Acetyltransferase; D005968:Glutamate Decarboxylase; C402565:glutamate decarboxylase 1; D002118:Calcium