Background: Several classifications of autism are currently accepted, however one specific distribution is divided into two groups, with high or low repetitive behaviors (Insistence on Sameness-IS). Twenty-five percent of the high IS group is also characterized as having high serotonin (5HT) in their platelets, while the rest having normal serotonin levels. Previous studies have shown that overall levels of 5HT receptor are unchanged so our hypothesis is that the differences in serotonin signaling exhibited by these patients are due to the changes in association between key signaling components. Serotonin, in lymphocytes, signal through 2 different G proteins with 5HT 4, 6, and 7 receptors coupling to Gαs leading to the activation of adenylyl cyclase, which can be attenuated when Gαs is sequestered in lipid rafts. Conversely, Gαq11, signaling through 5HT2A receptors, is enhanced when sequestered in lipid rafts. Changes in the localization of membrane-associated proteins in autism or ASD (autism spectrum disorder), downstream modifications could be taking place in regards to proper coupling between these 5HT receptors, their associated G proteins and signaling cascades. Methods: Lymphoblast cell lines, selected from subjects with high and low serotonin, were ordered from the National Institutes of Mental Health (NIMH) repository. Chronic effects of serotonergic drugs were studied with particular interest placed on the shift in membrane compartmentalization of important molecules for signaling including the G proteins: Gαs, Gαq11, and Gαi by Sequential TX100 (non-raft fraction) and TX114 (raft fraction) detergent extraction. Each was identified and quantified by immunoblotting. Signaling through serotonin 5HT4, 6 and 7 receptors (adenylyl cyclase) was investigated using a number of selective agonists and differences between cell lines were analyzed. Other cascades were probed as well including 5HT2 activation of Phospholipase C (PLC) and Adenylyl Cyclase inhibition through the 5HT1 receptor. Results: Membrane compartmentalization of G proteins, specifically Gαs, was shown between each different group with Gαs enriched in the non-raft membrane fraction of cells from hyperserotonemic subjects. Consistent with this (although in contrast to the expected depression of 5HT signaling in an elevated serotonin environment), 5HT4 or 5HT6 receptor activation was enhanced in the hyperserotonemic cell lines. There were no differences in Gαq11 compartmentalization between cells prepared from the two groups, and 5HT 2b response is currently being investigated. Discussion: Cells prepared from hyperserotonemic subjects show clear differences from normal serotonin subjects in lipid raft compartmentalization of Gαs. This translates into differences in serotonin signaling, particularly through 5 HT4 and 5 HT6 receptors. It’s important to point out that receptor number, specifically 5HT2A, does not change. Proper indication of responses to serotonergic drugs could point to narrowing down treatment options that would best fit individual patients and/or point to novel therapeutic mechanisms for autism.