Natural remedies are frequently used by people suffering from anxiety disorders, but evidence of their benefits in randomised controlled studies [30
] and laboratory research [18
] is limited. Due to the controversial nature of homeopathic claims, it is important for any results in this field to be confirmed and consolidated through further investigations and rigorous statistical evaluation. Two previous investigations [15
] suggested that Gelsemium s.
reduced anxiety and fear and increased exploratory behaviour in the laboratory mouse, without provoking any sedation side-effects. However, in the first series, the major and most significant effect was noted in OF parameters [15
], while, in the second one, the LD test yielded the best results [16
]. Since reproducibility, the degree of accordance between the results of experiments testing the same hypothesis, is a fundamental requisite for acceptance of any evidence, we performed a new analysis to evaluate statistically the differences between the two series and the global significance of the results.
In all parameters considered but one (distance travelled in centre in OF), there were no significant differences between the two experimental series nor interaction between series and experimental groups. This indicates that the trends of the drug effects were qualitatively in the same direction, despite a noteworthy quantitative variability. The pooled data analysis confirms and reinforces the evidence that statistically high significant Gelsemium s. effects can be detected in the laboratory mouse using both the OF and LD paradigms, even with the high dilutions/dynamizations employed in the homeopathic pharmacopoeia (9C and 30C). This laboratory evidence, based on blinded protocols and using groups of large sample size, strongly supports the conclusion that homeopathic medicaments are not mere placebos and are endowed with specific pharmacological activity.
The ability of extremely diluted drugs to change these emotional responses of mice can be ascribed to the high sensitivity of the tests involved, which are designed to put the animal in a situation of uncertainty (“bifurcation point,” indicated by an asterisk in ), where an extremely slight influence can determine the choice of which direction to move in (A or B in figure). The sensitivity of these tests to minimal factors is also, conceivably, one reason for the high variability of responses in the two series of experiments, observed in both vehicle-treated and drug-treated animals. It has been noted that the extent to which an anxiolytic compound facilitates exploratory activity depends on its baseline level in the control group [25
]. Bousta et al. [10
] report some anxiolytic-like effects of G. sempervirens
in mice stressed by repeated electric shocks, but no such effects in normal unstressed mice. Differences between the nature and severity of external stressors, or between experimental setups, environment, handling and testing, and individual biological responses to drugs, might account for the high variability of results reported under different experimental conditions [24
]. Variable behavioural baseline levels have been reported by others [17
], and it has been found that two groups having low and high “trait” anxiety and different neuroendocrine responses to stress can be selected from the same mouse population [35
], indicating that expression of trait anxiety displays a high interindividual variability in inbred mice.
In the OF model, Gelsemium s.-treated mice were unaffected in their general movement and locomotion in the field but showed a higher tendency to enter the central zone, instead of running along the walls or staying in the corners. This behaviour is thought to reflect changes in the emotional state of the mouse, even though in our experimental conditions, the OF parameters do not measure “anxiety” but rather exploratory propensity, thigmotaxis, and neophobia. This conclusion is based on the fact that neither buspirone nor diazepam altered those parameters. The differences between the effects of Gelsemium s. and those of the conventional anxiolytic drugs diazepam and buspirone suggest that the former has a broader action on animal behaviour, possibly including the stimulation of exploratory behaviour in the OF. The LD, on the other hand, proved to be a very valid test for anxiety, given that it always showed some effect with the two conventional anxiolytics, as well as with Gelsemium s.
Anxiety, neophobia, fear, and thigmotaxis are rather complex phenomena. There are two types of anxiety, “state” anxiety (excess anxiety experienced by a subject at a particular time in presence of a stimulus) and “trait” anxiety (does not vary from moment to moment) [36
]. It has been suggested that the light-dark test and elevated plus-maze device are the most appropriate models for assessing state anxiety, while the free-exploratory paradigm can be used for “trait anxiety” [33
]. It has also been reported that anxiolytic treatments do not by themselves increase exploration in the central zone of the OF, but they do decrease the stress-induced inhibition of exploratory behaviour [17
]. Benzodiazepines have been found to be inactive in some models or even to produce paradoxical anxiogenic effects [38
]. That OF is less sensitive to benzodiazepines, and buspirone as compared with other behavioural tests, (e.g., elevated plus-maze) has been shown also by others [39
], and a decrease of locomotion caused by buspirone at low (1
mg/kg) and high (10
mg/kg) doses has been observed in rats [40
]. Further studies with additional tests of anxiety are needed to confirm this intriguing relationship.
These findings strongly suggest that the LD and OF tests explore different emotional responses, with different sensitivities to drugs and neurological mechanisms. Our data showing lack of correlation between responses with two test used () seem in agreement with this conclusion. In this connection, it is also worth noting that the peak of Gelsemium s. activity in the LD test was observed with the 9C dilution dynamization, while, in the OF, it occurred with the 7C. This may suggest that the different behavioural “symptoms” exploited by these two test paradigms are sensitive to different dilutions/dynamizations of the remedy.
A possible action mechanism of Gelsemium s.
at neurological level has been indicated by others, showing that, in rat brain slices, very low doses [41
] and high dilutions/dynamizations (5C, 9C) of this compound [42
] enhance the production of the neurosteroid allopregnanolone (5a,3a-tetrahydroprogesterone), a stimulator of GABAa receptors and of inhibitory signalling in the central nervous system. These authors [41
] showed that this activity was stimulated by glycine and blocked by strychnine, well known as a glycine receptor (Gly-R) antagonist, suggesting that gelsemium effects are antagonistic to those of strychnine and mediated by Gly-R receptors.
is frequently used in homeopathy to treat patients exhibiting neurological anxiety-like symptoms such as “general prostration, trembling, tired feeling, mental apathy, muscular weakness, complete relaxation and prostration, lack of muscular co-ordination, general depression, emotional excitement, bad effects from fright, fear, exciting news” according to the Materia Medica [6
]. The fact that the traditional indications for the remedy are consistent with significant laboratory findings using rigorous experimental models helps bridge a gap between two medical disciplines generally considered to be at variance with each other, but which should instead be regarded as complementary and compatible. Of course, further scientific evidence of possible clinical benefits of homeopathy in humans is needed.