The Messinian salinity crisis (late Miocene) is one of the most fascinating paleoceanographic events in the recent geological history of the Mediterranean Sea, defining a time when it partly or nearly completely dried out. However, the relative roles of tectonic processes and sea-level changes, as triggers for restriction and isolation of the Mediterranean Sea from the open ocean, are still under debate. In this study, we present a detailed pollen, dinoflagellate cyst (dinocyst), and magnetic susceptibility analysis of a sequence of late Neogene (between ca. 7.3 and 5.2 Ma) marine sediments from the Montemayor-1 core (lower Guadalquivir Basin, southwestern Spain), which provides a continuous record of paleoenvironmental variations in the Atlantic side of the Betic corridors during the late Miocene. Our results show that significant paired vegetation and sea-level changes occurred during the Messinian, likely triggered by orbital-scale climate change. Important cooling events and corresponding glacio-eustatic sea-level drops are observed in this study at ca. 5.95 and 5.75 Ma, coinciding with the timing and duration of oxygen isotopic events TG32 and TG22–20 recorded in marine sediments worldwide. It is generally accepted that the onset of the Messinian salinity crisis began at ca. 5.96 ± 0.02 Ma. Therefore, this study suggests that the restriction of the Mediterranean could have been triggered, at least in part, by a strong glacio-eustatic sea-level drop linked to a climate cooling event occurring at the time of initiation of the Messinian salinity crisis.