Figure 14.

Workflow to screen for effects of cnidarian venom fractions on dorsal root ganglia (DRG) cells using constellation pharmacology. Primary cultures were plated, as described in the Section 5. Eighteen hours later, DRG cells were incubated with Fura-2 for 1 h (30 min in culture chamber and 30 min at room temperature). Continuous Fura-2 calcium imaging was conducted during pulse-chase exposure of the cells in the flow cell well to control solutions, depolarizing KCl pulses and the specific pharmacological agents comprising constellation pharmacology. Venom fractions were applied in between successive KCl pulses (blue box). Cell responses (Fura-2 ratiometric intensity over time) were transformed into profiles of individual cells to be analyzed. KCl, potassium chloride 25 mM; AITC, allyl isothiocyanate (agonist of the transient receptor potential ankyrin 1 channel or TRPA1); Men, menthol (agonist of the transient receptor potential melastatin 8 channel or TRPM8); Cap, capsaicin (agonist of the transient receptor potential vanilloid 1 channel or TRPV1).