A NOVEL PHARMACOLOGICAL APPROACH AND IDENTIFICATION OF PERIPHERAL CELLULAR BIOMARKERS IN NIEMANN-PICK C DISEASE PATIENTS

Aim of our project is to understand the contribute of lipid rafts in the occurrence of neurological disturbs in Niemann-Pick Disease type C (NPDc) and in particular their possible involvement in the incorrect development of neuronal networks. In fact, the loss of a correct dynamic of cholesterol-sphingolipids-enriched microdomains in the neuronal and glial plasma membrane, caused by an imbalance in the lipid trafficking due to NPDc gene mutation, could have a key role in neuronal dysfunction and the consequent clinical pathologies. Lipid rafts constituents can be manipulated with drugs either interacting with the metabolism of their components (i.e. inhibitors of the synthesis of glycosphingolipids or cholesterol), or extracting/adding single components by using liposomes or molecules that bind cholesterol (i.e. cyclodextrines). Although the genetic defects causing NPDc are well known, very few information is available on the causes of neurological symptoms that eventually lead to a fatal end the patient affected by NPDc. Changes in the functioning of neuronal plasma membranes are good candidates in the research of NPDc pathophysiogenetic mechanisms. Recently, it has been shown that neuronal mitochondria membranes are altered and unable to effectively participate in ATP synthesis, a defect that could be antagonized by normalizing cholesterol with methyl-b-cyclodextrin. Until now, no study addressed the possible changes in the functioning of neuronal plasma membrane receptors as a putative determinant of NPDc neurological disturbs. Changes in the plasma membrane cholesterol content and in the glycosphingolipids/cholesterol ratio is particularly important in affecting lipid rafts and neurotransmitter receptor activity. In particular, lipid rafts are important regulators of glutamate receptor functioning. Electrophysiological recordings and calcium imaging analysis have been performed on rodent hippocampal slices and cell cultures to study the effects of modification of lipid rafts composition both on synaptic transmission and on calcium influx in a control group of animals. We evaluated the effects of beta-cyclodextrine induced cholesterol depletion on synaptic transmission and on the expression of Paired Pulse Facilitation (PPF) in CA1 hippocampal region. We found that cyclodextrine strongly reduced synaptic transmission whereas it did not affect PPF. The role of glutamatergic receptors in cholesterol depletion mediated effects was evaluated by applying specific pharmacological agents. Our data indicate that, in cholesterol depleted neurons, reduction of synaptic transmission is sustained by AMPA and kainate receptors altered activities. Moreover, the involvement of these receptors has been confirmed by fluorimetric measurements of intracellular calcium concentration. These results support the hypothesis that modulation of receptors activity by manipulation of membrane lipids component may represent a possible therapeutical strategy in neurological aspects of NPDc