1 The analgesic properties of the synthetic cannabinoid WIN55,212-2 were investigated in a model of neuropathic pain. In male Wistar rats, bilateral hind limb withdrawal thresholds to cold, mechanical and noxious thermal stimuli were measured. Following this, unilateral L5 spinal nerve ligation was performed. Seven days later, sensory thresholds were reassessed and the development of allodynia to cold and mechanical stimuli and hyperalgesia to a noxious thermal stimulus con®rmed. 2 The eect of WIN55,212-2 (0.1 ± 5.0 mg kg 71 , i.p.) on the signs of neuropathy was then determined; there was a dose related reversal of all three signs of painful neuropathy at doses which did not generally alter sensory thresholds in the contralateral unligated limb. This eect was prevented by co-administration of the CB 1 receptor antagonist SR141716a, but not by coadministration of the CB 2 receptor antagonist SR144528, suggesting this action of WIN55,212-2 is mediated via the CB 1 receptor. Administration of SR141716a alone had no aect on the observed allodynia and hyperalgesia, which does not support the concept of an endogenous analgesic tone. 3 These data indicate that cannabinoids may have therapeutic potential in neuropathic pain, and that this eect is mediated through the CB 1 receptor.

Among the several mechanisms of multidrug resistance (MDR), overexpression of drug efflux pumps CaCdr1p and CaMdr1p belonging to ATP binding cassette (ABC) and major facilitator superfamily (MFS) respectively remain the predominant mechanisms of candidal infections. Therefore inhibiting or modulating the function of these transporters continues to draw attention as effective strategy to combat MDR. We have previously reported the antifungal potential of Geraniol (Ger), a natural monoterpenoid from Palmarosa oil, against Candida albicans. Herein, we explored the fungicidal nature of Ger. The Rhodamine 6G (R6G) and Nile red accumulation confirms the specific effect on CaCdr1p. Mechanistic insights with Candida cells overexpressing CaCdr1p and CaMdr1p revealed that Ger specifically modulates CaCdr1p activity. Kinetic studies further unraveled the competitive inhibition of Ger for R6G efflux as evident from increased apparent Km without affecting Vmax value. The effect of Ger on CaCdr1p was substantiated by molecular docking analyses, which depicted in-silico binding affinity of Ger with CaCdr1p and explored that Ger binds to the active site of CaCdr1p with higher binding energy. Although RT-PCR and western blot revealed no change in expressions of CDR1 and CaCdr1p, confocal microscopy images however depicted CaCdr1p mislocalization in presence of Ger. Interestingly, Ger was synergistic (FICI<0.5) with fluconazole (FLC) which is a well known antifungal drug. Furthermore, Ger sensitizes the FLC sensitive and resistant clinical matched pair of isolates Gu4/Gu5 and led to abrogated R6G efflux and depleted ergosterol. Furthermore, Rhodamine B labeling demonstrates altered mitochondrial potential with Ger which suggest possible linkage of dysfunctional mitochondria with CaCdr1p activity. We also estimated phenotypic virulence marker extracellular phospholipase activity which was considerably diminished along with inhibited cell adherence and biofilm biomass. Lastly, antifungal efficacy of Ger was demonstrated by enhanced survival of Caenorhabditis elegans model and negligible hemolytic activity (20%). Together, modulation of efflux pump activity by Ger and FLC synergism represent a promising approach for combinatorial treatment of candidiasis.

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