Isoform-specific induction of nuclear free calcium oscillations by platelet-derived growth factor

Confocal laser scanning microscopy was used to analyze alterations in nuclear free calcium (Ca²⁺(n)) levels induced by platelet-derived growth factor (PDGF) isoforms in BALB/c3T3 fibroblasts loaded with the calcium- sensitive fluorescent indicator Fluo-3. Both AA-PDGF and BB-PDGF caused a transient increase in Ca²⁺(n). Analysis of PDGF-induced Ca²⁺(n) alterations as a function of time revealed that BB-PDGF stimulation resulted in the generation of Ca²⁺(n) oscillations that diminished over time. The frequency of BB-PDGF-stimulated oscillations was modulated by extracellular Ca²⁺ and could not be mimicked by increasing intracellular inositol 1,4,5- trisphosphate levels in the absence of growth factor stimulation. Caffeine alone had no effect on Ca²⁺(n) levels, but exposure of cells to caffeine after BB-PDGF stimulation augmented Ca²⁺(n) oscillations, either by increasing the frequency or reinitiating preexisting oscillations. The genesis of these oscillations in Ca²⁺(n) appears to be in the region just outside of the nucleus, as perinuclear cytoplasmic free calcium (Ca²⁺(i)) increased just prior to Ca²⁺(n). In contrast, AA-PDGF stimulation resulted in the generation of one or two irregular, transient Ca²⁺(n) spikes. Caffeine pretreatment followed by AA-PDGF stimulation resulted in Ca²⁺(n) oscillations very similar to those produced by BB-PDGF alone. Additionally, the AA-PDGF and BB-PDGF isoforms appeared to modulate distinct pools of cellular Ca²⁺, as BB-PDGF was still capable of inducing Ca²⁺(n) oscillations subsequent to prior induction of oscillations by AA- PDGF/caffeine. These PDGF isoform-specific changes in nuclear free Ca²⁺ could serve as a mechanism by which isoform-specific cellular signaling pathways may be manifested by the growth factors.