Gelsemine and Gelsemium sempervirens L. Extracts in Animal Behavioral Test: Comments and Related Biases

due to Gelsemium noxious compounds if more selective compounds can be identified. Centesimal (C) dilutions of G. sempervirens (4C, 5C, 7C, 9C, and 30C) appeared able to change significantly mouse behavioral effects as assessed in the open-field (OF) and light–dark (LD) tests in blind and randomized fashion (Magnani et al., 2010) but effects in the elevated plus-maze, an assay considered to be important in behavioral assessment of potential anxiolytic compounds (Ramos, 2008), have not been assessed. Actually, the physical integration of different current tests in one single apparatus, in such a way that the emotional status of an animal becomes assessable through a series of distinct tasks, should contribute to increased reliability, rapidity, and comprehensiveness in behavioral testing. Although manuscripts in the literature have reported showing an anxiolytic effect of G. sempervirens at concentrations lower than 1.67 ymol/L, criticism may be raised about biases in their experimental context (Venard et al., 2009; Magnani et al., 2010). When a preparation according to the homeopathic pharmacopeia is assayed in a behavioral experimental setting with animals, the contribution of biases may overwhelm any reliable output (Bellavite et al., 2009). The main active substance in G. sempervirens causing an anxiolytic action is believed to be gelsemine (Bellavite, et al.,2009). Gelsemine, is an alkaloid that contains the same functional groups as strychnine and has been shown to increase 3α,5α-tetrahydroprogesterone or allopregnenolone (3α,5α-THP) neurosteroid synthesis (Venard et al., 2008). The stimulatory effects of glycine and gelsemine on 3α,5α-THP production were additive when the two drugs were combined. These results may suggest that glycine and gelsemine, acting via glycine receptor (GlyR), upregulate 3α,5α-THP biosynthesis in the spinal cord (Venard et al., 2008). Some pilot data have also raised the possibility that chronically administered neurosteroids antagonize certain acute effects of benzodiazepines and may enhance arousal via antagonist or inverse agonist actions at the benzodiazepine/GABA(A) receptor complex (Venard et al., 2008). As several complex issues were raised, the use of Gelsemium in behavioral science should undergo some reappraisal. In this field, a possible main critical topic might be the placebo/nocebo response of animals, able to exert biases on anxiety models. In medicine, the placebo response or placebo effect has often been regarded as a nuisance in basic research and particularly in clinical research, but very few works were related to animal behavior (McMillan, 1999). The latest scientific evidence has suggested, however, that the placebo effect and the nocebo effect, the negative effects of placebo, stem from highly active processes in the brain that are mediated by psychological mechanisms such as expectation and conditioning (Enck et al., 2008). Pavlovian conditioning might justify a placebo effect in housed laboratory animals (McDannald, 2010). Animals which undergo the same stressing procedure for several times (such as intra-peritoneal injections) may relate the physical stressor to other stimuli, such as operator’s odor or voice–sound, particularly when the same operator performs the injections. This bias is not always considered, although wash out and blinding randomized protocols are followed (Chirumbolo, 2011). The search for biases in animal behavior research should be a hallmark of any published paper on the topic. Some simple biases, for example animal sex (as researchers routinely use only male animals), have raised criticism concerning the opportunity to use these models to test drugs that would be used by the human female population (Wald and Wu, 2010). Placebo/nocebo response, as an effect of a conditioning/learning mechanism, is almost never considered in behavioral testing with laboratory animals: actually, none of the recently published papers about the Gelsemine and Gelsemium sempervirens L. extracts in animal behavioral test: comments and related biases