Fig. 4.
Pre-exposure of DRG neurons to the new A3AR agonist ICBM irreversibly blocks ICa and prevents the effect of a successive application of Cl-IB-MECA. A, C Experimental protocol used to compare the long-term effects of the prototypical A3AR agonist Cl-IB-MECA (30 nM) or the new, irreversible, A3AR agonist ICBM (1 µM) on ICa inhibition in DRG neurons. The protocol consisted of a 10-min exposure of DRG cultures to the A3AR agonist (added to the culture medium), followed by a 15-min washout (removal of the A3AR agonist-containing medium and perfusion of DRG neurons with the extracellular patch-clamp solution in the recording chamber) during which a stable baseline of peak ICa amplitude (peak ICa) was acquired by whole-cell patch-clamp recordings. Hence, the effect of a subsequent exposure to the prototypical A3AR agonist Cl-IB-MECA (30 nM) on ICa was assessed in the same cell. B, D Original current traces recorded in a Cl-IB-MECA (B) or in a ICBM (D) pre-incubated DRG neuron before and during the Cl-IB-MECA (30 nM) application. Scale bars: 500 pA; 100 ms. Lower panels: typical time course of ICa peak evoked in respective cells. Right panels: pooled data (mean ± SEM) of ICa peak measured before (baseline, bsl) or during the Cl-IB-MECA (30 nM, n = 5; B) or ICBM (1 µM, n = 7; D) application. P value refers to paired Student’s t-test