The present study used imaging and electrophysiological techniques in salamander retinal slices to correlate Ca²⁺ and Cl^- levels in rods and thus test the hypothesis of a feedback interaction between Ca²⁺ and Ca²⁺-activated Cl^- channels whereby Cl^- efflux through Cl^- channels can inhibit Ca²⁺ channels. Increasing [K⁺]_o levels produced a concentration-dependent depolarization of rods accompanied by increases in [Ca²⁺]_i measured with Fura-2. The voltage dependence of increases in [Ca²⁺]_i was compared to the voltage dependence of the calcium current (I_Ca). [Cl^-]_i was measured with the dye, MEQ. Depolarization with high K⁺ to membrane potentials below -20 mV reduced [Cl^-]_i; larger depolarizations increased [Cl^-]_i. The Na/K/Cl cotransport inhibitor, bumetanide, shifted the apparent Cl^- equilibrium potential (E_Cl) to more negative potentials suggesting that this cotransporter helps establish a relatively depolarized E_Cl. MEQ fluorescence changes evoked by high K⁺ were inhibited by niflumic acid (0.1 mM), NPPB (2 µM), or replacement of Ca²⁺ with Ba²⁺ suggesting that depolarization-evoked Cl^- changes result partly from stimulation of Ca²⁺-activated Cl^- channels. Replacing >12 mM [Cl^-]_o with CH₃SO₄^- produced a significant reduction in [Cl^-]_i. [Ca²⁺]_i increases evoked by 20 or 50 mM K⁺ were also significantly inhibited by replacing >12 mM [Cl^-]_o with CH₃SO₄^-. Thus, modest depolarization can evoke increases in [Ca²⁺]_i that lead to reductions in [Cl^-]_i and, conversely, reductions in [Cl^-]_i inhibit depolarization-evoked [Ca²⁺]_i increases. These findings support the hypothesis that feedback interactions between Ca²⁺ and Ca²⁺-activated Cl^- channels may contribute to the regulation of presynaptic Ca²⁺ currents involved in synaptic transmission from rod photoreceptors.