Data and calculus on isobolographic analysis to determine the antinociceptive interaction between calcium channel blocker and a TRPV1 blocker in acute pain model in mice

Determining antinociceptive interaction between Phα1β toxin (a voltage gated calcium channel blocker) and SB366791 (selective TRPV1 antagonist) may have both clinical and mechanistic implications for the pain management. This data in brief article is associated to the research paper “Synergistic antinociceptive effect of a calcium channel blocker and a TRPV1 blocker in an acute pain model in mice”. This material supports the isobolographic analysis performed with the above drugs and shows: data of the dose response curves of the agents given as single drug or combination regimens. Mathematics and statistical processing of dose response curves, proportion of drugs dosage to be used in the combination, calculus of theoretical additive DE20 dose as well as experimentally obtained DE20 are provided. It is also presented details of statistical comparison between theoretical and experimentally obtained DE20.


a b s t r a c t
Determining antinociceptive interaction between Phα1β toxin (a voltage gated calcium channel blocker) and SB366791 (selective TRPV1 antagonist) may have both clinical and mechanistic implications for the pain management. This data in brief article is associated to the research paper "Synergistic antinociceptive effect of a calcium channel blocker and a TRPV1 blocker in an acute pain model in mice". This material supports the isobolographic analysis performed with the above drugs and shows: data of the dose response curves of the agents given as single drug or combination regimens. Mathematics and statistical processing of dose response curves, proportion of drugs dosage to be used in the combination, calculus of theoretical additive DE 20 dose as well as experimentally obtained DE 20 are provided. It is also presented details of statistical comparison between theoretical and experimentally obtained DE 20  Isobolographic analysis using a fixed proportion ratio of two drugs.
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Data accessibility
Data is with this article.
Value of the data -Data tables and calculus from this article may serves as a practical guide on isobolographic analysis for testing other drug regimens of even using different pain models. -Determining the interaction index by isobolographic analysis provides a measure of the in vivo degree of interaction of two drugs for a specified effect. -Data analysis presented in this article can be systematically compared to other data from probit analysis with analgesic drugs given in combination.

Intrathecal Phα1β
See Table 3.   a ¼ ̅ y -b̅ x. thus b ¼ 12.9 (slope) and a ¼ 62    a ¼ ̅ y -b̅ x. thus b ¼ 21.4 (slope) and a ¼ 54  Table 7.  a ¼ ̅ y -b̅ x. thus b ¼ 20.1 (slope) and a ¼ 53 -Comparison test t critical ¼ (x-y)/[(SEx) 2 þ(SEy) 2 ] 1/2 ; wherein: ; where in: t add ¼ tabular value of t based on N´-2 degrees of freedom for 95% of significance t mix ¼ tabular value of t based on N-2 degrees of freedom for 95% of significance (SEy) 2 ¼ Variance of Z mix Interpretation: If t critical oT tabulated , then the difference is not significant, which means an additive effect. When t critical 4T tabulated , Z mix is significantly smaller (95% of confidence) than the Z add , which implies a synergistic effect of combined drugs (Table 8).

Capsaicin-induced nociceptive responses
This test was conducted essentially as described previously [1]. Briefly, individual animals were placed in transparent acrylic square boxes (20 cm per side) immediately after the capsaicin injection (1 nmol/paw i.p.), and nociceptive behaviours were recorded continuously for 300 s. Behaviours were quantified by recording the time spent licking, biting or flinching (nociceptive time) the paw injected with capsaicin. Phα1β (i.t. or i.p), SB366791 (i.p.), combined Phα1β þ SB366791 or their respective vehicles were injected 10 min before the capsaicin injection. The initial dose ranges of Phα1β and SB366791 were selected based on previous data [1]. Antinociceptive effects were measured as a percentage of the maximum possible (% MPE) effect according to the formula: % MPE ¼ 100*(A -B / A), where A is the averaged nociceptive time of the vehicle group and B is the nociceptive time of each animal in the treated group (drugs alone or in combination) [2].

Experimental design and statistical isobolographic analysis
Experimental design and statistical analyses were conducted essentially as previously described [3][4][5]. Briefly, dose-response curves were first obtained for Phα1β and SB366791 administered alone. Line equations, slope values and respective variances were obtained using linear regression [3]. DE 20 values and 95% confidence limits were calculated using probit analysis. DE 20 values (doses that exhibit 20% MPE) were used to assess whether the dose-effect of these drugs alone exhibited a constant potency ratio, which is necessary to perform fixed dose-pair combination of drugs [4]. Drug doses in association studies were determined as a proportion of their DE 20 values. This proportion was constant and estimated based on a factor derived from the individual variances of the DE 20 values. This fixed proportion of agents was necessary to assess whether the combination displayed enhanced potency indicative of synergism. Dose-response curves of associated drugs were constructed to obtain the doses that achieved the same effect level (20% MPE) compared to drugs given alone. This experimentally obtained DE 20 (here called Z mix ) was compared to theoretically calculated DE 20 value for additive interactions (Z add ). The criterion for establishing a statistical significance was P o 0.05. Graphical assessments of synergy are also presented using isobolographic analyses. Measurement of the interaction index (α) was obtained by dividing experimentally obtained DE 20 of the drug pair by the theoretical additive DE 20 of the drug pair. The α interaction index provides a measure of the degree of synergism.