Local and systemically delivered glucocorticoids are commonly administered to protect the cochlea against damage associated with a variety of insults. There is reason to believe that dexamethasone administered by these routes may arrive at cochlear target sites via different pathways. Clinically, there is a lack of clarity as to which route is more effective in any specific circumstance. This study explores dexamethasone distribution within the guinea pig cochlea following local and systemic delivery methods. A combination of mass spectroscopy and immunohistochemistry were employed to compare both perilymph distribution, tissue uptake and receptor activation. Local administration of dexamethasone to the round window membrane resulted in greater perilymph concentrations, with a basal to apical gradient that favours the cochlear base. Tissue immunofluorescence was intimately related to perilymph concentration following local administration. Systemic administration resulted in much lower perilymph concentrations, with an inverse basal to apical gradient favouring the cochlear apex. Lower perilymph concentrations following systemic administration were associated with minimal tissue immunofluorescence. Despite this, GR activation of the SGNs was equivalent in both administration regimes. These results bring into question the efficacy of measuring perilymph concentrations alone as a surrogacy for dexamethasone distribution and activity in the cochlea, suggesting that the steroid ligand may arrive at its target receptor via alternative pathways. Our results suggest an equivalence in efficacy between local and systemic administration routes early after drug delivery, when the ultimate outcome of GR activation is the goal.