Abstract
Background
Common clinical use of antipsychotics (AP) drugs shows that their therapeutic mode of action still needs further clarification although it is admitted that the Dopamine receptor D2 (D2R) antagonism plays a significant role. For instance, clozapine (CLOZ) - which is known to be the most effective AP in treating schizophrenic symptoms - has strikingly the lowest D2R antagonism. Non direct receptor-related effects might thus be involved in the activity of AP at the synapse level.
AP, as well as neurotransmitters, are mostly lipophilic and insert within membranes. This characteristic is of interest as a significant proportion of schizophrenic patients has specific and abnormal membrane lipid composition. This possible proxy of the disease biotype can participate in the disease’s physiopathology but also be critical for the effect of AP drugs.
We hypothesize that AP insertion into lipid membranes also contribute to their therapeutic effect. AP-induced modifications of synaptic membranes biophysics are likely to influence neurotransmission. In this study, we focus on the effect of AP on membrane fusion, a crucial step for the exocytosis of neurotransmitters.
Methods
Liposomes modelling synaptic vesicles were reconstituted in saline buffer. Two standard ternary and quaternary lipid mixtures have been studied: phosphatidylcholine:phosphatidylethanolamine:phosphatidylserine (PC:PE:PS) [65:25:10] and the synaptic-like PC:PE:PS:sphingomyelin:cholesterol (PC:PE:PS:SM:CHOL) [25:25:10:10:30]. Some liposomes were protein-free and others were functionalized with Soluble N-ethylmaleimide-sensitive-factor Attachment protein Receptor (SNARE) proteins, which trigger in vivo the fusion of synaptic vesicles with the pre-synaptic plasma membrane. The liposome size was checked by Dynamic Light Scattering. Insertion of AP within the membrane was checked by second derivative spectroscopy. Fusion was measured by Fluorescence Resonance Energy Transfer in the absence or presence of CLOZ or chlorpromazine (CPZ) at various lipid:AP ratios (10:1 to 100000:1). Protein-free liposomes were fused with Polyethylene glycol (PEG) and SNARE liposomes through the action of cognate SNARE proteins residing in their membrane.
Results
Liposomes of the same lipid composition were of the same size, with no effect of the addition of AP drugs at various concentrations. Molar partition coefficient of AP drugs within the membrane of protein-free liposomes was approximately 70–85%. CPZ or CLOZ inhibited the fusion of PC:PE:PS liposomes by about 20–40%. When liposomes were synaptic-like (PC:PE:PS:SM:CHOL), the inhibition of fusion by AP drugs reached 50%. CLOZ also inhibited SNARE-mediated fusion of PC:PE:PS liposomes by about 30%. This effect on SNARE-mediated fusion was not observed with CPZ.
Discussion
Altogether, these results, despite preliminary, could help to understand partially a non direct receptor-related effect of antipsychotics. Indeed, these drugs also seem to modify membrane dynamics at the synapse level. This seems to be particularly the case of CLOZ on SNARE-mediated fusion and could explain its specific therapeutic efficiency.
The Msb3/Gyp3 GAP controls the activity of the Rab GTPases Vps21 and Ypt7 at endosomes and vacuoles
