Gallino, S. | bioRxiv Science

bioRxiv2022

Cannabinoids Activate the Insulin Pathway to Modulate Mobilization of Cholesterol in C. elegans

The nematode Caenorhabditis elegans requires exogenous cholesterol to survive and its depletion leads to early development arrest. Thus, tight regulation of cholesterol storage and distribution within the organism is critical. Previously, we demonstrated that the endocannabinoid (eCB) 2-arachidonoylglycerol (2-AG) plays a key role in C. elegans modulating sterol mobilization, but the mechanism is unknown. Here we show that mutations in the ocr-2 and osm-9 genes coding for transient receptors potential V (TRPV) ion channels, dramatically reduces the effect of 2-AG in cholesterol mobilization. Through genetic analysis combined with the rescuing of larval arrest induced by sterol starvation we found that the insulin/IGF-1signaling (IIS) pathway and UNC-31/CAPS, a calcium-activated regulator of neural dense-core vesicles release, are essential for 2-AG-mediated stimulation of cholesterol mobilization. These findings indicate that 2-AG-dependent cholesterol trafficking requires the release of insulin peptides and signaling through the DAF-2 insulin receptor. These results suggest that 2-AG acts as an endogenous modulator of TRPV signal transduction to control intracellular sterol traffic through modulation of the IGF-1 signaling pathway. Author summaryAlthough cannabis extracts have been used in folklore medicine for centuries, the past few years have seen an increased interest in the medicinal uses of cannabinoids, the bioactive components of the cannabis plant, for treatment of many diseases of the nervous system. However, the human body naturally produces endocannabinoids that are similar to the cannabinoids present in Cannabis sativa. Our goal is to understand how endocannabinoids maintain cholesterol homeostasis in animals, underscoring the importance of cholesterol balance for healthy life. Both cholesterol excess and cholesterol deficiency can have detrimental effects on health, and a myriad of regulatory processes have thus evolved to control the metabolic pathways of sterol metabolism. The nematode C. elegans is auxotroph for sterols, that is; contrary to mammals they cannot synthesize sterols, therefore, dietary supply is essential for survival. The aim of our study was to elucidate the mechanism by which endocannabinoids abolish larval arrest of C. elegans induced by cholesterol depletion. We discovered that endocannabinoids stimulate the insulin pathway, which affects development, reproduction and life span, to modulate mobilization of cholesterol in C. elegans. Our studies have important implications for a better understanding of human pathological conditions associated with impaired cholesterol homeostasis.

neuroscience↗

bioRxiv2020

Loss of choline agonism in the inner ear hair cell nicotinic acetylcholine receptor linked to the α10 subunit

The 910 nicotinic acetylcholine receptor (nAChR) plays a fundamental role in inner ear physiology. It mediates synaptic transmission between efferent olivocochlear fibers that descend from the brainstem and hair cells of the auditory sensory epithelium. The 9 and 10 subunits have undergone a distinct evolutionary history within the family of nAChRs. Predominantly in mammalian vertebrates, the 910 receptor has accumulated changes at the protein level that may ultimately relate to the evolutionary history of the mammalian hearing organ. In the present work we investigated the responses of 910 nAChRs to choline, the metabolite of acetylcholine degradation at the synaptic cleft. Whereas choline is a full agonist of chicken 910 receptors it is a partial agonist of the rat receptor. Making use of the expression of 910 heterologous receptors, encompassing wild-type, heteromeric, homomeric, mutant, chimeric and hybrid receptors, and in silico molecular docking, we establish that the mammalian (rat) 10 nAChR subunit underscores the reduced efficacy of choline. Moreover, we show that whereas the complementary face of the 10 subunit does not play an important role in the activation of the receptor by ACh, it is strictly required for choline responses. Thus, we propose that the evolutionary changes acquired in the mammalian 910 nAChR resulted in the loss of choline acting as a full agonist at the efferent synapse, without affecting the triggering of ACh responses. This may have accompanied the fine-tuning of hair cell post-synaptic responses to the high frequency activity of efferent medial olivocochlear fibers that modulate the cochlear amplifier. Contribution to the Field StatementIn the inner ear of mammals, several evolutionary changes have occurred resulting in an expansion of the hearing range to higher sound frequencies. Fine tuning of cochlear synapses is required for sound enconding. The synapse between efferent olivocochlear fibers, that descend from the hindbrain, and sensory hair cells modulates sound amplification at the periphery and it has been proposed as a major player in the expansion of the hearing range. The 910 nicotinic acetylcholine receptor, which mediates synaptic neurotransmission at the efferent fiber-hair cell synapses, has accumulated a high number of amino acid substitutions in the mammalian lineage. We now show that these evolutionary acquired changes have led to a mammalian receptor with a lower efficacy for choline, the metabolite produced at the synaptic cleft by acetylcholine degradation. Making use of molecular, electrophysiological and in silico simulations techniques we show that it is the 10 subunit the one responsible for the loss of full choline agonism on the efferent receptor in mammals. This functional change may prove fundamental to faithfully reproduce the high frequency activity of efferent medial olivocochlear fibers and the modulation of the cochlear amplifier.

neuroscience↗