Calcium signaling in a narrow somatic submembrane shell during synaptic activity in cerebellar Purkinje neurons

Temporal and spatial changes in the intracellular Ca²⁺ concentration ([Ca²⁺](i)) were examined in dendrites and somata of rat cerebellar Purkinje neurons by combining whole-cell patch-clamp recording and fast confocal laser-scanning microscopy. In cells loaded via the patch pipette with the high-affinity Ca²⁺ indicator Calcium Green-1 (K(d) ≃ 220 nM), a single synaptic climbing fiber response, a so-called complex spike, resulted in a transient elevation of [Ca²⁺](i) that showed distinct differences among various subcellular compartments. With conventional imaging, the Ca²⁺ signals were prominent in the dendrites and almost absent in the soma. Confocal recordings from the somatic region, however, revealed sleep transient increases in [Ca²⁺](i) that were confined to a submembrane shell of 2- to 3-μm thickness. In the central parts of the soma [C²⁺](i) increases were much slower and had smaller amplitudes. The kinetics and amplitudes of the changes in [Ca²⁺](i) were analyzed in more detail by using the fast, low-affinity Ca²⁺ indicator Calcium Green-5N (K(d) ≃ 17 μM). We found that brief depolarizing pulses produced [Ca²⁺](i) increases in a narrow somatic submembrane shell that resembled those seen in the dendrites. These results provide direct experimental evidence that the surface-to-volume ratio is a critical determinant of the spatiotemporal pattern of Ca²⁺ signals evoked by synaptic activity in neurons.