Intracellular calcium spiking generates versatile signals that control various key neuronal functions. However, estimation of neuronal spiking pattern at subcellular spatial resolution remains challenging. In this context, we propose calcium imaging to estimate activities in soma, dendrite and axon, along with neuronal interconnections and network topology. Accordingly, we develop a novel integrated framework for neuronal analysis consisting of experimental and computational techniques. Specifically, we present a combination of high resolution imaging technique using confocal microscopy, neural signal processing involving deconvolution, and model-based simulation of neuronal calcium spiking. Importantly, we validate aspects of the proposed simulation with experimental results for hippocampal neuron populations. Our technique has a unique advantage over more ubiquitous methods, such as electrophysiological studies, in the ability to provide the micro-organization of the neuronal network and corresponding spike patterns at subcellular spatial resolution. © 2017 IEEE.