Journal: The Journal of General Physiology
Article Title: Distinct purinergic signaling pathways in prepubescent mouse spermatogonia
Figure Lengend Snippet: P2X4 is a candidate mediator of the high-affinity ATP response in spermatogonia. (A) Reverse transcription PCR reveals transcripts for P2rx2 , P2rx4 , and P2rx7 in both Sertoli–germ cell co-cultures and whole testis preparations at P7. A faint band for P2rx3 -encoding cDNA is only amplified from whole testis samples but not co-cultures. Primer pairs generate amplificates of 150–250 bp, respectively. cDNA from whole brain/spinal cord serves as positive control, whereas no template controls are devoid of detectable signals. (B–E) Exemplary whole-cell voltage-clamp recordings of ATP-induced currents (10 µM; 1 s; V hold = −80 mV; S 1 , S 6 ) under control conditions and during treatment with suramin (10 µM and 100 µM; B), extracellular Cu 2+ (100 µM; C), reduced pH (6.3; D), and ivermectin (3 µM; E). Altered conditions were established for at least 60 s (preincubation). (F) Bar diagram (means ± SEM) depicting the quantitative analysis of experiments exemplified in B–E. Note the discontinuous ordinate (//). Numbers of cells tested are indicated above bars. Asterisks (*) denote statistical significance, P ≤ 0.01 (paired t test).
Article Snippet: Primary antibodies were anti-DAZL (1:500; Abcam), anti-Sloα1 (extracellular; 1:500; Alomone Labs), anti-P2X4 (extracellular; 1:100, Alomone Labs), and anti-P2X7 (extracellular; 1:500; Alomone Labs).
Techniques: Polymerase Chain Reaction, Amplification, Positive Control