By 2035, bees, responsible for up to 75% of food consumed, may disappear in the United States if the current rate of population decline continues, with declines strongly associated with pesticide exposure (Benjamin & McCallum, 2008). Neuropesticides, including neonicotinoids and fipronil, act on areas of the brain and lead to altered behaviors, often supported by neurodegenerative processes and physiological modifications (Rortais et al., 2005). In response to this global problem, and after extensive evidence, certain neonicotinoids, along with fipronil, were banned in Europe (Kathage et al., 2018). However, concerns against a widespread market ban point to the fact that farmers may rely on older pesticides, potentially more toxic or susceptible to evolving pest resistance, leading to higher costs of food (Özkara et al., 2016). Therefore, approaches in the near future must guarantee food security considering the conservation of crops and pollinators (van der Sluijs & Vaage, 2016). Here we evaluate a neuropharmacological protection strategy provided by plant-derived secondary metabolites against fipronil, a neuropesticide widely used in many places around the world (Simon-Delso et al., 2015). Honey bees and bumblebees are considered key pollinators around the world; However, other wild bees also face similar threats (Klein et al., 2018; Potts et al., 2016). In Apis mellifera scutellata and Bombus impatiens, sublethal doses of fipronil impair cognitive functions, reducing individual and colony performance (Frazier et al., 2015; van der Sluijs & Vaage, 2016). In the brain of bees, fipronil targets areas such as the Mushroom Bodies (MBs), centers underlying learning, memory among other cognitive functions (Decourtye et al., 2009; El Hassani et al., 2005, 2009; Jacob et al., 2015). Within MBs, neurodegeneration of microglomeruli (MGs; subregions that exhibit rich neuronal connectivity and plasticity) and biochemical changes in neuronal ATP levels presumably explain cognitive and physiological deficits (Cintra-Socolowski et al., 2016 ; Nicodemo et al., 2014; Therefore, sublethal exposures to fipronil decrease key skills, such as navigation and resource evaluation (Decourtye et al., 2009; Pisa et al., 2015). Consequently, individual deficiencies of bees affect the higher level of organization and colonies can collapse (Simon-Delso et al., 2015; Steinhauer et al., 2018). In this context, pharmacologically protecting bees from the negative effects of fipronil is a key approach that directly impacts their health and supports food security. In this work, we have discovered that the administration of several plant-derived secondary metabolites, such as rutin flavonols, kaempferol and p-coumaric acid (a phenolic acid), successfully protects cognitive and neurostructural processes against sublethal fipronil exposure. Furthermore, we have shown that sublethal doses of fipronil and imidacloprid, two different classes of neuropesticides, not only impair the cognitive performance of bees, but also alter and reduce mitochondrial ATP production. Therefore, based on our findings, we propose that the aforementioned phytochemicals could protect at the physiological and mitochondrial level in honey bees and/or bumblebees that are exposed to sublethal doses of fipronil and imidacloprid. B. impatiens and A. mellifera foraging bees prophylactically treated with rutin, kaempferol, p-coumaric acid or a mixture of these, and subsequently exposed to a sublethal dose of fipronil chronically or acutely, had learning protection and protection at a neurostructural level, which did not differ from unexposed bees. On the contrary, and as reported in the literature, bees exposed to fipronil exhibited a significant impairment in learning, memory, and mitochondrial ATP production El Hassani et al., 2009; Nicodemo et al., 2014; Riveros & Gronenberg, 2022). Therefore, it is crucial to identify the level of action and mechanisms supporting the protection supported by phytochemicals. Investigation of these aspects will support the use and specific design of doses, and will provide greater evidence of physiological and neurostructural protection, as well as allow the evaluation of additional phytochemicals that have been studied in the context of neurodegenerative diseases (Kumar & Khanum, 2012; Nkpaa and Onyeso, 2018). Here we investigate protection at several levels: behavioral (learning and memory), neurostructural (neurodegeneration/neuroprotection of MGs) and we investigate the impairment caused by two different neuropesticides at the physiological (production of mitochondrial ATP levels) and cognitive (learning and memory) levels. Our research was based on the honey bee A. mellifera and the bumblebee B. impatiens due to their relevance as the main pollinators and due to their key advantages as classical experimental models (Matsumoto et al., 2012; Riveros & Gronenberg, 2009). Finally, this research not only contributes to the understanding of the mechanisms associated with the protection of bees, but also establishes a solid foundation for future studies aimed at their conservation.
