This study evaluates the potential water availability in Barka Slough and the effects of changing hydrological conditions on the aquatic habitat of five protected species. Barka Slough is a historically perennial wetland at the downstream western end of the San Antonio Creek Valley watershed (SACVW). A previously published hydrologic model of the SACVW for 1948–2018 was extended to include 2019–2021 and then modified to simulate the future years of 2022–2051. Two models simulating the future years of 2022–2051 were constructed, each with different climate inputs: (1) a repeated historical climate and (2) a 2070-centered Drier Extreme Warming climate (2070 DEW). The model with the 2070 DEW climate had warmer temperatures and an increase in average annual precipitation driven by larger, albeit more infrequent, precipitation events than the model with the historical climate. Simulated groundwater pumpage resulted in cumulative groundwater storage depletion and groundwater-level decline in Barka Slough in both future models. The simulations indicate that Barka Slough may transition from a perennial to an ephemeral wetland. Streamflow, stream disconnection, and depth to groundwater are key habitat metrics for federally listed species in Barka Slough. Future seasonal conditions for each metric are more likely to affect federally listed species’ habitats under 2070 DEW climatic conditions. Future seasonal streamflow volume may negatively impact unarmored threespine stickleback (Gasterosteus aculeatus williamsoni) and tidewater goby (Eucyclogobis newberryi) habitats. Future seasonal stream disconnection may negatively impact the unarmored threespine stickleback habitat. Future groundwater-level decline may negatively impact Gambel’s watercress (Nasturtium gambelii) and La Graciosa thistle (Cirsium scariosum var. loncholepis) habitats and could influence the ability to use Barka Slough as a restoration or reintroduction site for these species. Results from this study can be used to inform water management decisions to sustain future groundwater availability in the SACVW.