Rxfp2a and Rxfp2b mediate Insl3 action in the zebrafish testis.A Effects of zebrafish Insl3 on four relaxin family peptide receptors (Rxfps) expressed in HEK293T cells transiently transfected with pcDNA3.1 and pCRE plasmids. Data are expressed as mean ± SEM (N = 3, technical replicates). Numbers in brackets indicate the EC50 concentrations. AU, arbitrary units. B Expression levels of responsive Insl3 receptors in control, germ cell-depleted (by exposure to the cytostatic agent busulfan51), and testes with recovering (from busulfan) spermatogenesis, as described by Crespo et al.25 (NCBI GEO data set GSE116611). Data are expressed as mean ± SEM (N = 5; *p < 0.05). C, D Localization of rxfp2a:EGFP (green; C) and rxfp2b:mCherry signal (red; D) in adult testis tissue. Confocal laser scanning microscopy analysis of whole-mount testes shows preferential expression of EGFP and mCherry in type A spermatogonia and Sertoli and myoid cells, respectively. DAPI counterstain is shown in gray. Representative germ cell types (insets 1–4) and Sertoli and myoid cells are shown (insets 5–6). Aund, type A undifferentiated spermatogonia; Adiff, type A differentiating spermatogonia; B, type B spermatogonia; SPC, spermatocytes. In D, red and white arrowheads indicate representative mCherry+ and mCherry− somatic cells, respectively. Scale bars, 25 µm.
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