2.1.3. Drug administration
A u-tube system (Fig 1) was used for the rapid application of capsaicin.
The u-tube was placed close to a group of cells of interest, and this
was attached to a gravity-feed perfusion system through microbore Teflon
PTFE tubing (O.D. 0.042”, I.D. 0.022” (Cole Palmer). The tubing had
previously been coated with a siliconising agent (sigmacote,
Sigma-Aldrich), and the application of capsaicin was controlled by a
3-way solenoid valve (The Lee Company, England) connected to pulse
generator unit (High Med, England). The drug application lasted 10s for
all experiments involving capsaicin.
capsaicin was applied twice with a 10 min rest period, at the second
application the magnitude of the calcium responses was diminished
compared to the response obtained following the first capsaicin
application (mean ± sem) response ratio which is described as the (F510
self-ratio) of the second response as a percentage of the first response
(F510 self-ratio).
The different chemical reagents used to inhibit the sensitisation effect
of bradykinin on capsaicin cells were started 10–30 min before the
challenge with the first capsaicin and continued until the experiment
done. Each chemical was dissolved in the appropriate solvent at the
desired concentration, according to the manufacturer’s recommendations.
All reagents were diluted to the desired concentration in a normal
Ringer’s solution (pH 7.4) before the experiments (table1).
Table 1: The
concentrations and solvents applied to investigate their role in
capsaicin sensitization by bradykinin.