Our results indicated that fluoxetine altered resting [Ca2+]ER, partially, which may be correlated with ER stress, that has emerged as an important event in the pathophysiology of diabetes. release and store-operated calcium access (SOCE) activation, probably through reduction of ER calcium storage and inhibition of stromal conversation molecule 1 (STIM1) trafficking. These data suggest that exposure to fluoxetine results in impaired cell functions, occurring in concert with reduction of E-cadherin-dependent cell adhesion and alterations of calcium homeostasis. Introduction Patients with major depressive disorder (MDD) have a higher incidence of type 2 diabetes mellitus (T2DM) when compared to the general populace1, 2. Even though underlying mechanism(s) involved in the relationship between T2DM and MDD is not fully understood, recently a growing number of studies indicate that long-term use of SSRIs constitutes to a major risk factor for impaired glucose homeostasis and development of T2D3C5. Similarly, a recently population-based, nested case-control study in Taiwan showed a 20% increased risk of diabetes for patients with long-term antidepressant treatment for two or more years6. Despite these findings, little is known about the direct pathophysiology of SSRIs on pancreatic cell functions. Early studies exhibited that administration of fluoxetine and fluvoxamine induced hyperglycemia in rodents7, 8. Isaac model32. Cells were incubated with fluoxetine, a widely used SSRIs33, for 3?h. Our results showed that fluoxetine (30?M) had no effect on cell proliferation WS 12 and cell viability (Fig.?S1A,B); however, it significantly inhibited GSIS (Fig.?S1C). Next, we sought to understand the cellular and molecular events underlying this deleterious effect of fluoxetine on insulin secretion. Cell-cell adhesion plays an important role in regulating GSIS from pancreatic cells16, 18, so next we examined whether fluoxetine can affect cell morphology, and cell-cell adhesion. Our results showed that MIN6 cells grew in tightly packed colonies with close cell-cell contact in the control group, while cells created smaller colonies of WS 12 loosely packed cells with reduced cell-cell contact in the fluoxetine-treated group (Fig.?1A). To assess the role of adhesion molecules in mediating the alteration in cell morphology, MIN6 cells were immuno-stained with Alexa 488 (green) for E-cadherin and Alexa 594 (reddish) for -catenin (Fig.?1B). We found control group with adjacent cells within each colony shared common boundaries demarcated by E-cadherin, but E-cadherin was reduced at area of cell contact and cell dispersed after fluoxetine treatment (Fig.?1B). Here we defined three characteristics of cell populations from our confocal images by performed z-section from top to bottom of cells (Fig.?1C). Combined cells stood for cells stick together at each stage, while separated cells represented that cells were totally disconnected from the top to bottom. Interestingly, there were some cells being associated to each other at the middle stage, but separated at the top and bottom stage. We defined this populace as semi-separated cells. Quantification of these three characteristics of cell populations from confocal images stage-by-stage, as shown in Fig.?1D, 96.1??2.7% of control cells combined to other cells, but only 67.2??8.6% of fluoxetine-treated cells remained combined. The results indicated that fluoxetine altered cell morphology correlated with a loss of cell-cell adhesion. Open in a separate window Physique 1 Fluoxetine alters cell morphology, and reduces cell-cell adhesion. (A) After 3-hour fluoxetine (30?M) treatment, MIN6 cells were observed under an inverted fluorescence microscope (Evos). The white arrows show reduction of cell-cell adhesion. Level bar, 100?m. The representative images were from at least three impartial experiments. (B) After 3-hour incubation with or without fluoxetine (30?M), MIN6 cells were fixed and then immuno-stained with Alexa 488 (green) for E-cadherin, Alexa 594 (red) for -catenin and EPOR Hoechst 33258 (blue) for nucleus. The images were captured by using confocal microscope (Olympus, MPE). Level bar, 10?m. The representative images were from at least three impartial experiments. (C) Schematic diagram defines three characteristics of cell contact. Cells were categorized by how close they contact to each other WS 12 at different z-sections. Cell junction was fully continuous from top to bottom (combined cells), partially lost at the top and bottom (semi-separated cells) or totally lost from top to bottom (separated cells). (D) Quantitative analysis for the percentage of each contact type of MIN6 cells treated with or without fluoxetine. Each value represents imply??SEM of at least 600 individual cells. Fluoxetine alters WS 12 the structure of adherens junction and the distribution of E-cadherin Adherens junction is the most important structure to maintain cell-cell adhesion. E-cadherin connects neighboring cells at outer membrane34, and is regulated by cytosolic protein -catenin35, 36..