DISCUSSION - AXIOLYTIC PROPERTY OF CAESALPINIA PULCHERRIMA LEAVES EXTRACTS IN EXPERIMENTAL ANIMALS

 

DISCUSSION

 

            Anxiety and depressive disorders are common in all religions of the world.¹³⁶ They constitute a substantial proportion of the global burden of disease and are projected to form the second most common cause of disability by 2020.¹³⁷ This increased importance of non-communicable diseases such as anxiety and depressive disorders.¹³⁸

 

            Benzodiazepines are the major class of compounds that are used in anxiety and they remain the most commonly prescribed treatment for anxiety. However, the realization that BZD have a narrow safety margin has prompted many researchers to evaluate new compounds in the hope of identifying other anxiolytic drugs with fewer side effects.¹³⁹

 

            The use of herbal medications by physicians in Europe and Asia is becoming very common and researchers are exploring the traditional remedies to find a suitable cure for these ‘mind affecting diseases’.¹⁴⁰ The role of herbal medicine in the treatment of various psychological disorders has become well established over the past two decades. Many herbal products used to treat anxiety, cognitive disorders and insomnia contain chemicals that influence the function of ionotropic receptors for the brain’s major inhibitory neurotransmitters GABA, serotonin and BZD.¹⁴¹

 

            Despite the widespread traditional use of Caesalpinia pulcherrima  for treating various disorders there are no reports of scientific evaluation of its anxiolytic activity, therefore the present study was undertaken to explore anxiolytic potential of the leaves of Caesalpinia pulcherrima  using anxiety models like, Elavated plus maze, Hole board test, Light/Dark transition test and Open field test. Acute toxicity study was conducted for ethanolic and petroleum ether extracts and after 21 days study it was concluded that both the extracts were safe upto 2000 mg/kg body weight with fewer side effects like grooming and hair loss and no adverse effects.

 

            The elevated plus maze is currently one of the most widely used model of animal anxiety and it’s validated and use in rats and mice⁹⁴. The EPM test is based on a premise where the exposure to an EPM evoked an approach-avoidance conflict that was considerably stronger than evoked by the exposure to an enclosed arm⁸⁹. The decrease in aversion to the open arm is the result of an anxiolytic effect, expressed by the increased time spent and entries in to the open arm are sensitive to agents thought to act via the GABAA receptor complex, justifying the use of diazepam as a positive control in this study⁹¹, even when the compound being screened does not act via benzodiazepine receptors.

 

            Diazepam has increased the percentage of entries in open arms (***P < 0.001) and time spent in open arms (**p< 0.01), the percentage of entries in close arms (***P< 0.001), and percentage of time spent in closed arms (*P<0.05) has decreased significantly compared to control group. It was seen that the ethanolic extract (400 mg/kg) and petroleum ether extract (400 mg/kg) are statistically non-significant in all the parameters of EPM test. While ethanolic extract (800mg/kg) has significantly increased the percentage of open arm entry (*p < 0.01) and time spent in open arms (*p < 0.05) and petroleum ether extract (800 mg/kg) has also increased the percentage of open arm entry (**p < 0.01) and time spent in open arms (**p < 0.01) more significantly than ethanolic extract when compared to control group.

            The hole-board test provides a simple method for measuring the response of an animal to an unfamiliar environment and is widely used to assess emotionality, anxiety and/or responses to stress in animals. The head dipping behavior was sensitive to changes in the emotional state of the animal, and suggested that the expression of the anxiolytic state in animals may be reflected by an increase in head dipping behavior¹⁴².

 

            Diazepam has showed significant increase in all the parameters of HBT. It was found moderately significant in head dip count and no. of assisted rearing             (**p< 0.01), while it was more significant in head dip latency and locomotion     (***p< 0.001). The ethanolic extract (400 mg/kg) and petroleum ether extract (400 mg/kg) were found to be statistically non-significant in all the parameters of HBT. Ethanolic extract (800 mg/kg) was found moderately significant in head dip latency and locomotion (**p< 0.01) and less significant in head dip count and no. of assisted rearing (*p< 0.05). Whereas petroleum ether extract (800 mg/kg) is more significant in head dip count, head dip latency and locomotion (**p< 0.01) and less significant in no. of assisted rearing (*p< 0.05) when compared to control group.

 

            Light/dark box is another widely used rodent anxiety model for screening anxiolytic or anxiogenic drugs. It is based on the innate aversion of rodents to brightly illuminated areas and on the spontaneous exploratory behavior of rodents in response to mild stressors that is novel environment and light¹³². Drugs induced increase in behavior in the white part of a two compartment box, in which a large white compartment is illuminated and a small black compartment is darkened, is suggested as an index of anxiolytic activity¹³³.

            Diazepam and both the extracts (ethanolic 800mg/kg and petroleum ether 800mg/kg) have more significantly increased the time spent in dark area            (***p< 0.001) and decreased the time spent in light area more significantly          (***p< 0.001) when compared to control group. While ethanolic extract (400 mg/kg) and petroleum ether extract (400 mg/kg) have increased the time spent in dark area less significantly and the time spent in light area was decreased less significantly    (*p< 0.01).

 

            The Open Field Test (Walsh & Cummins, 1976) provides simultaneous measures of locomotion, exploration and anxiety. The number of line crosses and the frequency of rearing are usually used as measures of locomotor activity, but are also measures of 4 exploration and anxiety. A high frequency of these behaviours indicates increased locomotion and exploration and/or a lower level of anxiety. The number of central square entries and the duration of time spent in the central square are measures of exploratory behaviour and anxiety. A high frequency/duration of these behaviours indicates high exploratory behaviour and low anxiety levels.¹⁴³

 

            In the open field test diazepam and both the extracts at a dose of 800mg/kg increased the locomotion and no. of assisted rearing more significantly (***p< 0.001). And entries in the central square and time spent in central square increased were statistically less significant (**p< 0.01) when compared to control group. While ethanolic and petroleum ether extracts at a dose of 400mg/kg were non-significant in all the parameters.

 

            Earlier reports on the chemical constituents of plants and their pharmacology suggests that plants containing flavonoids, alkaloids, phenolic acids, essential oils, saponins and tannins possess activity against many CNS disorders.¹⁴⁴ Investigations on the phytochemical screening of Caesalpinia pulcherrima leaves extracts revealed the presence of glycosides, saponins, tannins, flavonoids and monoterpenoids like, linalool (Table 2). From the literature survey it was also found that Caesalpinia pulcherrima leaves contains flavonoids like sitosterol and quercetin which have been reported earlier to possess anxiolytic activity.

 

            Thus, in this study both ethanolic and petroleum ether extracts of Caesalpinia pulcherrima leaves at a dose of 800mg/kg and diazepam (2mg/kg) significantly possessed anxiolytic property in a dose dependent manner (400; 800mg/kg) in all the parameters of anxiety models mentioned, when compared to control group.