A simple and fast densitometric method for the analysis of tyrosine hydroxylase immunoreactivity in the substantia nigra pars compacta and in the ventral tegmental area.
Parkinson's disease is a progressive dyskinetic disorder caused by degeneration of mesencephalic dopaminergic neurons in the substantia nigra pars compacta (SNpc) and, to a lesser extent, in the ventral tegmental area (VTA). Tyrosine hydroxylase (TH) is a rate-limiting enzyme for dopamine synthesis, therefore immunohistochemistry for TH can be used as an important marker of dopaminergic cell loss in these regions. Traditionally, immunohistochemical experiments are analyzed qualitatively by optical microscopic observation or more rarely semi-quantitatively evaluated by densitometry. A common problem with such papers is the lack of a clear explanation of the algorithms and macros employed in the semi-quantitative approaches. In this paper, we describe, in detail, an easy, fast and precise protocol for the analysis of TH immunoreactivity in SNpc and VTA using one of the most popular image analysis software packages (Image Pro-Plus). We believe that this protocol will facilitate the evaluation of mesencephalic TH immunoreactivity in various available animal models of Parkinson's disease.
Xavier
Léder Leal
LL
Departamento de Ciências Fisiológicas, Faculdade de Biociências, Pontifícia Universidade Católica do Rio Grande do Sul, Rua Ipiranga 6681, CEP 90619-900, Porto Alegre, RS, Brazil. llxavier@pucrs.br
Viola
Giordano Gubert
GG
Ferraz
Anete Curte
AC
Da Cunha
Claudio
C
Deonizio
Janyana Marcela Doro
JM
Netto
Carlos Alexandre
CA
Achaval
Matilde
M
D000818:Animals; D002138:Calibration; D002452:Cell Count; D003720:Densitometry; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D016627:Oxidopamine; D051381:Rats; D017208:Rats, Wistar; D013378:Substantia Nigra / Q000187:drug effects / Q000201:enzymology*; D013563:Sympathectomy, Chemical; D013565:Sympatholytics; D016707:Tissue Fixation; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*; D017557:Ventral Tegmental Area / Q000187:drug effects / Q000201:enzymology*
D007202:Indicators and Reagents; D013565:Sympatholytics; D016627:Oxidopamine; D014446:Tyrosine 3-Monooxygenase