title: A novel use of combined tyrosine hydroxylase and silver nucleolar staining to determine the effects of a unilateral intrastriatal 6-hydroxydopamine lesion in the substantia nigra: a stereological study.
abstract: Neurotoxic lesions of the nigrostriatal pathway model the deficits found in Parkinson's disease. This study used stereology and a novel staining method to examine the effects of a partial unilateral striatal 6-hydroxydopamine (6-OHDA) lesion on substantia nigra pars compacta (SNpc) dopamine neuron number and morphology in rats. Adult male Long-Evans rats were subjected to unilateral lesion of the SNpc by intrastriatal microinjection of 6-OHDA (12.5 μg). Lesions were verified by d-amphetamine-stimulated rotation (2.5 mg/kg, sc) by force-plate rotometry 7 days post-surgery. Seven days after rotation testing, rats were euthanized, and brains were prepared for either histology (n=12) or determination of striatal dopamine content by HPLC-EC (n=20). Brains prepared for histology were stained for tyrosine hydroxylase (TH) combined with a silver nucleolar (AgNOR) stain using a modified protocol developed for stereological assessment. The AgNOR counterstain allowed for precise definition of the nucleolus of the cells, facilitating both counting and qualitative morphometry of TH-positive neurons. Stereological quantitation determined a 54% decrease in TH-positive neuron number (P<0.01), and a 14% decrease in neuron volume (P<0.05) on the lesioned side. Striatal dopamine concentration was decreased by 92% (P<0.01), suggesting that striatal dopamine analysis may overestimate the numbers of SNpc neurons lost. These findings demonstrate that combined use of TH and AgNOR staining provides improved characterization of 6-OHDA-induced pathology. Furthermore, the data suggest that decreased neuronal volume as well as number contributes to the functional deficits observed after unilateral intrastriatal 6-OHDA lesion.
authors: Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, USA.
authors: Michelle
authors: M
authors: Healy-Stoffel
authors: S Omar
authors: SO
authors: Ahmad
authors: John A
authors: JA
authors: Stanford
authors: Beth
authors: B
authors: Levant
mesh_terms: D018663:Adrenergic Agents / Q000633:toxicity*; D000661:Amphetamine / Q000494:pharmacology; D000818:Animals; D003342:Corpus Striatum / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D004195:Disease Models, Animal; D004298:Dopamine / Q000378:metabolism; D007839:Functional Laterality / Q000187:drug effects / Q000502:physiology*; D008297:Male; D009474:Neurons / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D020258:Neurotoxicity Syndromes / Q000209:etiology* / Q000473:pathology / Q000503:physiopathology; D016627:Oxidopamine / Q000633:toxicity*; D051381:Rats; D020318:Rats, Long-Evans; D012399:Rotation; D016622:Silver Staining / Q000379:methods; D013238:Stereotaxic Techniques; D013378:Substantia Nigra / Q000378:metabolism / Q000473:pathology*; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*
chemical_list: D018663:Adrenergic Agents; D016627:Oxidopamine; D000661:Amphetamine; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine