title: Functional autoradiography and gene expression analysis applied to the characterization of the alpha2-adrenergic system in the chicken brain.
abstract: Here we report a functional autoradiographic study of [(35)S]GTPgammaS binding induced by alpha(2)-adrenoceptor activation in chicken brain tissue sections using both 10(-4)M UK 14304 (bromoxidine or brimonidine) and 10(-6)M epinephrine as alpha(2)-adrenoceptor agonists. Assays were performed using two different incubation buffers: glycylglycine or Tris-HCl. Changes in the [(35)S]GTPgammaS basal binding values were detected, and different [(35)S]GTPgammaS specific binding values were also obtained depending on the buffer used for each drug. The best results were obtained with epinephrine in Tris-HCl, with slightly higher stimulation values than the observed with UK 14304 in glycylglycine buffer. The effect of the addition of adenosine deaminase to the incubation buffer was also tested. This effect decreasing basal binding in chicken was very small when compared to mammals, according with differences found in adenosine 1 receptor expression levels. Structures presenting alpha(2)-adrenoceptor-mediated G(i/o) protein stimulation fitted with areas previously described as enriched in alpha(2)-adrenoceptors in chicken brain, and their homologous areas in mammals. These data confirm the specificity of the results and reinforce the implication of the alpha(2)-adrenoceptors in the function of these brain nuclei. On the other hand, the expression level of the different alpha(2)-adrenoceptor subtypes was tested with real-time PCR. Contrasting with the alpha(2)-adrenoceptor subtype distribution previously described with radioligand competition assays, where alpha(2A) was the predominant alpha(2)-adrenoceptor subtype (>/=75%); in the present work, the ratio of alpha(2A):alpha(2B/C) gene expression was lower than expected both in telencephalon, tectum opticum, and cerebellum.
authors.affiliation: Centro de Investigación Biomédica en Red de Salud Mental, CIBERSAM, Spain.
authors.forename: Rebeca
authors.initials: R
authors.lastname: Diez-Alarcia
authors.forename: Ricardo
authors.initials: R
authors.lastname: Mostany
authors.forename: Severiano
authors.initials: S
authors.lastname: Dos-Anjos
authors.forename: Arsenio
authors.initials: A
authors.lastname: Fernández-López
mesh_terms: D000243:Adenosine Deaminase / Q000378:metabolism / Q000494:pharmacology; D000316:Adrenergic alpha-Agonists / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography; D001667:Binding, Competitive / Q000187:drug effects / Q000502:physiology; D001921:Brain / Q000033:anatomy & histology / Q000378:metabolism*; D001931:Brain Mapping; D002395:Catecholamines / Q000378:metabolism*; D002645:Chickens / Q000033:anatomy & histology / Q000235:genetics* / Q000378:metabolism*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008297:Male; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D018341:Receptors, Adrenergic, alpha-2 / Q000235:genetics* / Q000378:metabolism*; D018047:Receptors, Purinergic P1 / Q000187:drug effects / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013045:Species Specificity; D013462:Sulfur Radioisotopes
chemical_list: D000316:Adrenergic alpha-Agonists; D002395:Catecholamines; D021122:Protein Subunits; D018341:Receptors, Adrenergic, alpha-2; D018047:Receptors, Purinergic P1; D013462:Sulfur Radioisotopes; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000243:Adenosine Deaminase; D019206:GTP-Binding Protein alpha Subunits, Gi-Go