We used local microclimatic conditions and twig sap flow rates to interpret midday stomatal closure in the canopies of two 250-year-old Norway spruce (Picea abies (L.) Karst.) trees at a subalpine site in the Swiss Alps (1,650 m a.s.l.). Both trees showed midday stomatal closure on most clear summer days, despite the permanently wet soil. We used a modified Penman-Monteith formula to simulate potential transpiration of single twigs (ET(T)) based on high-resolution temporal and spatial microclimate data obtained both inside and outside the crowns. Comparison of calculated ET(T) values and measured twig sap flow rates enabled us to pinpoint the occurrence of midday stomatal closure and the microclimatic conditions present at that time. We found that vapor pressure deficit (and for upper-crown twigs, ET(T)) largely explained the timing of initial midday stomatal closure but gave no explanation for the different patterns of stomatal behavior after initial closure in upper- and lower-crown twigs. After the initial stomatal closure, upper-crown twigs maintained high transpiration rates by continuously regulating stomatal aperture, whereas stomatal aperture decreased rapidly in lower-crown twigs and did not increase later in the day. Midday stomatal closure in lower-crown twigs occurred on average 1 h later than in upper-crown twigs. However, the microclimate at the time of initial stomatal closure was similar at both crown locations except that lower-crown twigs received significantly less solar radiation than upper-crown twigs both at the time of initial stomatal closure and afterwards. High rates of sap flow in twigs did not always lead to stomatal closure and therefore could not explain the phenomenon. We conclude that stomatal conductance can be modeled accurately only when both local microclimatic conditions and tree water status are known. Further, we hypothesize that both the quantity and quality of light play an important role in the reopening of closed stomata during the day.