ABSTRACT The secretion of insulin and glucagon by pancreatic β and α cells, respectively, is critical for glucose homeostasis. While the insulin granule dynamics are well‐characterized, the intracellular behavior of glucagon secretory granules (GSG) remains poorly understood. Here, we analyze the mobility of GSGs in αTC1‐9 cells and insulin secretory granules (ISG) in INS‐1E cells using spatiotemporal correlation spectroscopy and single‐particle tracking (SPT), with a focus on the role of the cytoskeleton in regulating their transport. Under basal conditions, SPT classification reveals that GSGs predominantly exhibit diffusive motion (57.6% ± 10%), with smaller fractions categorized as almost immobile (35.8% ± 10.6%) or drifted (6.6% ± 3%), closely resembling ISG dynamics. By disrupting microtubules, we confirmed their role as active tracks for directed granule transport in both cell types. Upon exposure to their respective secretory stimuli—high glucose for β cells and low glucose for α cells—both granule populations underwent a comparable shift toward increased diffusive and drifted motions. Treatment with the actin depolymerizing agent Latrunculin‐B reproduced this stimulatory effect in INS‐1E cells but not in αTC1‐9 cells, suggesting that despite their overall similarity in granule behavior under physiological conditions, α and β cells may rely on partially distinct mechanisms to engage the cytoskeletal network.