Discipline: Biological Sciences
Subcategory: Physiology and Health
Ahmed Hoda - California State University, Los Angeles
Co-Author(s): Li Andrew, Darabe Narek, Zhou Feimeng, Limon Agenor, and Russo-Neustadt Amelia, California State University, Los Angeles, CA
Recent scientific evidence suggests that hyperexcitability in neural circuits may be responsible for neurodegeneration and cognitive impairment during aging. This may, in turn, be due to an imbalance between excitation and inhibition neurotransmission or a reduction in GABAergic signaling. Thus, developing a way to prevent hyperexcitability in the brain can be an excellent strategy for preventing neuronal death and cognitive decline. Withania somnifera (WS), also known as ashwagandha, is herbal medicine that is been used for centuries in Ayurvedic medicine and is known as an adaptogenic plant because of its stress reducing capability. We have chosen to study this plant because evidence indicates that it has calming and GABA-enhancing capabilities. Our preliminary studies show that whole root WS modulates hippocampal activity and survival via a GABA-mimetic mechanism. In addition, the most widely studied active components of WS root extract, withaferin A and withanolide A, were not responsible for the observed GABA receptor activation. Thus, we hypothesize that other key WS constituents promote optimal hippocampal function and neuronal survival via GABAergic mechanism. To test this hypothesis, whole WS extracts have been separated into eight fractions using liquid-liquid extraction. To identify bioactive fractions with GABA activity, Xenopus laevis oocyte model was utilized to study GABArho (GABAA subtype) receptor function. Cloned GABArho cDNA from rat brain was injected into Xenopus oocyte. Two days after injections, GABA currents were recorded from voltage clamped oocytes using microelectrodes filled with 3M KCl and membrane potential held constant at -80 mV. The oocytes were continuously perfused with ringer solution at room temperature, followed by perfusion of the eight extracted WS fractions at 1: 100 dilution to test their physiological activity. Results showed that two out of eight fractions (water and butanol) exhibit significant maximal GABA response as compared to 10 uM GABA as a control (One Way ANOVA, p < 0.0001). This finding supports our hypothesis that major WS constituents may be responsible for the GABA-mimetic activity and have specificity for GABArho receptors as shown by significant GABA response in the two identified WS fractions and minimal response in the remaining six WS fractions. These results suggest that these constituents may be polar substances such as withanosides IV and VI, or small molecules such as amino acids. Understanding the mechanism of WS may be valuable for the development of pharmacological treatments for neurological disorders associated with GABAergic signaling dysfunction.
Funder Acknowledgement(s): I thank Zhou Feimeng for helping with liquid-liquid partitioning of WS. Agenor Limon for helping with electrophysiology set up. Funding was provided by NSF grant # HRD-0802628.
Faculty Advisor: Amelia Russo- Neustadt, ARusson@exchange.calstatela.edu
Role: I planned and performed the voltage clamp recording using the Xenopus oocyte model to study the effect of WS constituents on GABA receptors. To do this, I prepared the oocyte and a series of dilutions of WS liquid-liquid fractions , recorded current flow, and statistically analyzed the results using one way ANOVA.