Postsynaptic calcium influx at single synaptic contacts between pyramidal neurons and bitufted interneurons in layer 2/3 of rat neocortex is enhanced by backpropagating action potentials

Pyramidal neurons in layer 2/3 (L2/3) of the rat somatosensory cortex excite somatostatin-positive inhibitory bitufted interneurons located in the same cortical layer via glutamatergic synapses. A rise in volume-averaged dendritic [Ca2+]i evoked by backpropagating action potentials (APs) reduces glutamatergic excitation via a retrograde signal, presumably dendritic GABA. To measure the rise in local [Ca2+]i at synaptic contacts during suprathreshold excitation, we identified single synaptic contacts in the acute slice preparation in pairs of pyramidal and bitufted cells each loaded with a Ca2+ indicator dye. Repetitive APs (10-15 APs at 50 Hz) evoked in a L2/3 pyramidal neuron gave rise to facilitating unitary EPSPs in the bitufted cell. Subthreshold EPSPs evoked a transient rise in [Ca2+]i of 80-250 nm peak amplitude at the postsynaptic dendritic site. The local postsynaptic [Ca2+]i transient was restricted to ∼10 μm of dendritic length, lasted for ∼200 msec, and was mediated predominantly by NMDA receptor channels. When EPSPs were suprathreshold, the evoked AP backpropagated into the apical and basal dendritic arbor and increased the local [Ca2+]i transient at active contacts by approximately twofold, with a peak amplitude reaching 130-450 nm. This value is in the range of the half-maximal dendritic [Ca2+]i, evoking retrograde inhibition of glutamate release from boutons of pyramids. The localized enhancement of dendritic Ca2+ influx at synaptic contacts by synaptically evoked backpropagating APs could represent one mechanism by which a retrograde signal can limit the excitation of bitufted interneurons by L2/3 pyramids when these are repetitively active. ![Figure 5.][1] Figure 5. Presynaptic and postsynaptic Ca2+ influx at single synaptic contacts recorded by using two-photon excitation fluorescence microscopy. A , top, Fluorescence image of a pyramid-to-bitufted cell pair filled with rhod-2 and OGB-1, respectively. The pyramidal neuron is shown in red, the interneuron in green. The region with an identified synaptic contact is marked by the rectangle and shown at higher magnification at the bottom. The arrowhead points to a synaptic contact. The presynaptic axon is hardly visible, but lightens up during Ca2+ influx (see B ). Line scans shown in B were performed at the region indicated by the white vertical line. B , Fluorescence line scans recorded during unitary synaptic stimulation on the presynaptic bouton (red) and the postsynaptic dendrite (green). In the pyramidal neuron three APs were evoked at 10 Hz (see C ). C , Each of the three APs evoked in the pyramidal neuron gave rise to a Ca2+ influx into the presynaptic terminal (top traces, same experiment shown in B ). The postsynaptic neuron responded with two EPSPs, but only during one was a postsynaptic Ca2+ signal also measured (lower traces). Presumably, the second EPSP was evoked at another contact. Fluorescence data were not filtered. Note: No Ca2+ transient was recorded on the other dendrite also covered by the line scan (see B ), indicating the synaptic origin of the Ca2+ signals. [1]: pending:yes