The predicted increase in allochthonous dissolved organic carbon (DOC) in high-altitude lakes is expected to alter the phytoplankton–bacterioplankton relationship. However, few studies address the influence of summer phytoplankton dynamics on microbial responses to DOC additions. We sampled natural plankton assemblages during two contrasting periods of summer in a high-altitude lake in the French Alps and subjected them to glucose and nutrient enrichments under two light conditions (dark or light) and two temperature conditions (10°C or 18°C). Our results indicate that glucose use by bacteria differs over the summer, depending on the availability of autochthonous DOC and the nutrient limitation. Glucose was consumed by bacteria more in early summer; however, biomass increased with glucose addition more in late summer than in early summer. This pattern arose from the greater availability of phytoplankton-derived DOC in late summer, reducing the need for alternative carbon sources in late summer, when phytoplankton biomass was high. Mixotrophic taxa were stimulated after glucose additions both in early summer and in late summer. We found greater competition between bacteria and phytoplankton in late summer after glucose addition, linked to the summer nutrient limitation pattern. Our study thus highlights a differential response depending on the timing of summer DOC inputs. The global changes forecast for the French Alps should increase heterotrophic and mixotrophic processes in planktonic communities of shallow high-altitude lakes with vegetated catchments. This experimental study provides insights that will be useful in predicting ecological trajectories and in refining predictions of sentinel lakes' responses to global changes.