Large conductance, calcium-activated potassium (BK) channels play a central role in modulating cell excitability and contractility in vascular smooth muscle (VSM). BK channels are composed of pore-forming α subunits and accessory β subunits. The β subunits influence the calcium sensitivity of BK currents and in VSM the BKβ1 subunit is preferentially expressed. During the course of this meeting we show that in diabetes, the BKβ1 subunit is downregulated in retinal VSM cells. Work from other groups has suggested that high glucose can activate the transcription factor, NFATc3 (Nilsson et al. 2006). Activation of NFATc3 signalling is responsible for downregulating BKβ1 expression during angiotensin II-induced hypertension (Nieves-Cintron et al, 2006). Here we show that high glucose downregulates BKβ1 subunit expression in the A7r5 VSM cell line and that this is associated with nuclear accumulation of NFATc3. A7r5 cells were cultured in DMEM, with 2% calf serum in normal glucose (5 mM D-glucose) or high glucose (25 mM D-glucose) for times ranging between 30 min to 2 weeks. mRNA and protein expression for the β1 subunit was determined by real-time PCR and flow cytometry, respectively. Immunohistochemistry was used to assess NFATc3 activity in cells counter-stained with propidium iodide nuclear stain. For scoring of NFATc3 positive nuclei 30 fields of view were imaged for each time point, and an average of 50 cells per field were counted. All experiments were performed in triplicate. After 6 hours exposure to high glucose, β1 transcript levels decreased by approximately 40%, and this level of downregulation persisted up to 2 week’s exposure (p<0.01). This was paralleled by a 25% reduction in immunodetected β1 protein which was first evident 16 hours after high glucose treatment and was also sustained throughout the 2 week period (p0.05). Exposure of A7r5 cells to high glucose increased NFATc3 nuclear accumulation in a time-dependent manner. The number of NFAT positive nuclei increased from 21 ± 3% in control cells to 37 ± 2% and 47 ± 3% in cells exposed to high glucose for 30 min and 2 hours, respectively (p<0.001). These results suggest that A7r5 cells may represent a useful model system for investigating the molecular mechanisms underlying changes in BKβ1 expression in vascular myocytes during high glucose exposure, and furthermore, highlights the possible contribution of NFATc3.