Voltage-gated Ca2+ (Cav) channels are activated upon depolarization. They specifically allow Ca2+ ions to come into the cell. These Ca2+ ions are bi-functional because they not only control cell excitability but also couple electrical activity to complex downstream signaling events, such as excitation-contraction coupling in muscles and neurotransmitter release in neurons. In the brain, Cav channels are expressed in the pre- or post-synaptic membrane of most excitable cells, neurons. In the past few years, their expression and function have also been characterized in many nonexcitable cells such as astrocytes. This dissertation focuses on the regulation of one subtype of postsynaptic Cav channels, Cav1.2, in neurons. In the first part of chapter I, I provide a literature overview of Cav channels in terms of their subtypes, localizations, physiological functions, and biophysical properties. For years, Cav channels were studied as single entities. But now, based on multiple proteomic studies, we know that these channels actually do not live alone. They interact with numerous proteins depending on the physiological conditions. Such interactions can anchor the channels to optimal sites of action, and tether Cav channels to their modulatory molecules. Therefore, it is crucial to understand how Cav channels are regulated by their macromolecular assembly. Among these protein partners, our lab studied the regulation of Cav channels by a subset of PDZ-domain containing proteins. Because these proteins play an important role in scaffolding and they colocalize with both pre- and post-synaptic Cav channels. Indeed, previous studies from our lab and other groups have revealed that PDZ proteins participate in a multitude of Cav regulation. The second part of chapter I introduces the diverse modulation of neuronal Cav channels by numerous PDZ proteins. In neurons, Cav1.2 channels regulate neuronal excitability and synaptic plasticity. Their functions have been implicated in learning, memory, and mood regulation. A study published in the journal Lancet showed that the gene encoding Cav1.2 is a common risk factor for five major psychiatric disorders. A PDZ protein, densin-180 (densin) is an excitatory synapse protein that promotes Ca2+-dependent facilitation of voltage-gated Cav1.3 Ca2+ channels in transfected cells. Mice lacking densin exhibit similar behavioral phenotypes that closely match those in mice lacking Cav1.2. In chapter II and III, we investigated the functional impact of densin on Cav1.2 channels and their auxiliary subunit β2a. Besides the regulation of Cav channels by their interactome, we have also known for a long time that Ca2+ currents undergo a negative feedback regulation. This regulation is called Ca2+-dependent inactivation (CDI) and it is mediated by Ca2+ that directly traverses the pore. CDI has been described for Cav channels in multiple cell types. In the heart, CDI prevents excessively long cardiac action potentials, which in turn can prevent activity-dependent arrhythmia. In neurons, CDI may be neuroprotective by preventing excitotoxic Ca2+ overloads. In the last 18 years, two essential components have been revealed in the mechanism of CDI. One is the protein calmodulin (CaM). CaM interacts directly with sites on the C-terminus of Cav channels. It binds to the incoming Ca2+ ions and produces a mysterious conformational change that determines the conductance of the channel. The other molecular player is Cavβ protein family. Cavβ comprises four subfamilies β1 through β4, which generally enhance the channel inactivation, except β2a. In chapter IV, Xiaohan Wang from Roger Colbran’s lab in Vanderbilt University, and I identified a new molecular determinant for Cav1.2 CDI. The α2δ subunit is an extracellular component of the Cav channel complex. Similar to Cavβ subunits, α2δ subunits are essential for the biophysical properties, surface level, and trafficking of Cavα1 subunits. There are four isoforms of α2δ subunits (α2δ1 to α2δ4). They display distinct tissue distributions. Although the roles of α2δ subunits in Cav channel regulation were studied extensively, studies have proposed that the function of α2δ subunits may be in part or entirely independent of Cav channel complex, such as synaptogenesis. Considering the important role of α2δ in physiology and pathology, it is imperative to identify the factors that regulate the properties of α2δ. In chapter V, I explored the trafficking dynamics of α2δ1 and revealed a potential regulator of α2δ1 for its protein stability and localization. One beauty of doing research is that it always motivates us to think and ask more questions on our journey of demystifying nature. While looking at the evidence that I find, I realize how much more we could do in the future. In chapter VI, I conclude the findings of each chapter and share my perspectives on the future direction for these research projects.