0157 A BRAIN-SPECIFIC LIPID TRANSPORTER MODULATES NEURONAL AND BEHAVIORAL AROUSAL

Abstract

Phosphatidylinositol transfer proteins (PITPs) regulate phosphoinositide metabolism and play diverse roles in multicellular organisms, from cancer regulation to sensory neuron signal transduction. One class II PITP family member, Piptnc1, has been implicated in cancer metastasis yet is also expressed in the developing and adult vertebrate brain. Its endogenous function is unknown. We have discovered that zebrafish mutants that lack Piptnc1a are behaviourally hyperactive due to the hyper-activation of neurons implicated in arousal and found small molecule inhibitors capable of rescuing these phenotypes. Using Crispr/Cas9 mediated gene editing, we deleted the zebrafish ortholog piptnc1a. To discover neurological phenotypes, we then video monitored piptnc1a mutants and wild type sibling behaviour. We examined neuronal activity patterns via expression of immediate early genes and brain-wide changes in phosphorylated ERK, a readout of neuronal activity. Morphing these maps onto reference brains identified discretely affected neuronal populations. Finally, we discovered drugs capable of rescuing the neuronal and behavioural mutant phenotypes by testing the piptnc1a mutants’ responses to a dose series of small molecules. Compared to wild type and heterozygous siblings, piptnc1a mutants are behaviourally hyperactive across the 24 hour day/night cycle. Consistent with the behaviour, piptnc1a mutants show a dramatic upregulation c-fos as well as elevated P-ERK levels in several arousing nuclei, including the hypocretin/orexin neurons. In vitro characterization of mammalian PIPTNC1 function revealed that knockdown of PIPTNC1 levels led to a strong induction of the Pi3 kinase-AKT signal transduction pathway. Remarkably, at doses with no discernible effect on wild type siblings, both Pi3K and AKT small molecule inhibitors return piptnc1a mutant c-fos levels to baseline and also rescues mutant behavioural hyperactivity. We conclude that piptnc1a is required to maintain proper signalling via the Pi3K-AKT pathway in neurons involved in vertebrate arousal. When this pathway is upregulated, neurons that increase arousal are hyper-activated, leading to an improper setpoint for behavioural activity.