Simultaneous recordings of membrane voltage and concentration of intracellular Ca2+ ([Ca2+]i) were made in apical dendrites of layer 5 pyramidal cells of rat neocortex after filling dendrites with the fluorescent Ca2+ indicator Calcium Green-1. Subthreshold excitatory postsynaptic potentials (EP-SPs), mediated by the activation of glutamate receptor channels, caused a brief increase in dendritic [Ca2+]i. This rise in dendritic [Ca2+]i was mediated by the opening of low-voltage-activated Ca2+ channels in the dendritic membrane. The results provide direct evidence that dendrites do not function as passive cables even at low-frequency synaptic activity; rather, a single subthreshold EPSP changes the dendritic membrane conductance by opening Ca2+ channels and generating a [Ca2+]i transient that may propagate towards the soma. The activation of these Ca2+ channels at a low-voltage threshold is likely to influence the way in which dendritic EPSPs contribute to the electrical activity of the neuron.