Exocytosis elicited by ACh was increased by varenicline, nicotine or by both agonists together in rat chromaffin cells
The final goal of the use of rat chromaffin cells was to investigate whether the chronic treatment of the cells with nicotine, that will mimic the situation of a chronic smoker, will increase exocytosis. In a latter step, we investigated whether the effects of these drugs on exocytosis could be due to an effect on the nicotinic currents. First, we investigated whether previous results obtained in Fig. 1 were reproducible in rat chromaffin cells. We performed the same protocol described in Fig. 1 in chromaffin cells of ten rats. Representative recordings under the control condition, varenicline, nicotine or both drugs together are shown in Fig. 3A-D, respectively. Exocytosis elicited under control conditions amounted to 107.5 ± 30.1 fF (n=21). The same protocol was tested with 100 nM varenicline or 250 nM nicotine, eliciting 173.6 ± 40.6 fF (n=18) and 171.8 ± 49.2 fF (n=16) of exocytosis respectively. Varenicline and nicotine evoked 196.5 ± 38.9 fF (n=20, *p) of exocytosis. There was a period of 5 min wash-out between the different treatments. Average values are shown in Fig. 3E.
In order to compare these responses and due to the high variability of the control responses (from 24 fF to 466 fF), each result was compared and normalized with its own control. Varenicline increased the response to 2.2 ± 0.4 fold (n=18; *p) and nicotine increased it to 2.4 ± 0.6 fold (n=16; *p). Both drugs perfused together increased by 2.9 ± 0.5 fold the exocytotic response (n=20; *p) (Fig. 3F). There was no change in the membrane potential when the different drugs were perfused during the Step 2. Under control conditions, the membrane potential was -56.4 ±2 mV (n=21), and it reached the values of -57.3 ± 2.7 mV (n=18), -57 ± 2.6 mV (n=16) and -58 ± 2 mV (n=20) in the presence of varenicline, nicotine and varenicline perfused together with nicotine, respectively.