The general aim of the Olcese Laboratory's research is to define the role of ion channels in the control and regulation of cell excitability. Ion channels are membrane proteins that regulate the flow of ions across the cell membrane that in turn control a variety of functions such as neuronal excitability, heartbeat, muscle contraction and hormones release. Ion channels are transmembrane proteins usually associated with accessory subunits that modulate their function. The regulation mechanism(s) of voltage-dependent channel activity by accessory subunits and second messengers is one of our strongest interests. In these studies, by combining state-of-the-art electrophysiological techniques (cut-open oocyte voltage clamp, patch clamp, macropatch) with mutagenesis, we are identifying the structural determinants for the various channel functions (activation, inactivation, conduction and sensitivity to drugs).
Voltage-dependent ion channels respond to changes in membrane potential by a rearrangement of the voltage sensing structure. The movement of the voltage sensor produces a conformational change of the channel that allows ion conduction through the pore. Part of our research is currently dedicated to understanding the coupling mechanism between the movements of the voltage sensor and pore opening. This work involves the measurement of small currents (gating currents) generated by the movement of the charged voltage sensor across the transmembrane electric field. Gating currents reveal the conformational changes of the channel protein, occurring before and during channel opening. In this research program, we combine optical methods with classical electrophysiology to detect conformational changes of the channels. This novel approach in ion channel studies allows the detection of conformational alterations of channel protein as changes in the fluorescence properties of fluorophores covalently attached to selected positions on the protein (voltage clamp + site-directed fluorescence labeling). One of the goals is to correlate at the molecular level the structural changes of the channel protein with the different phases of channel activity. We are using this approach to investigate the properties of voltage- and calcium-activated (BK) channels. The final objective is to obtain a molecular and dynamic view of BK channels by identifying the mobile regions underlying voltage sensing.
We are currently investigating how the amino acids in close proximity of the conduction pore are affecting the conduction properties of voltage- and calcium-activated K+ channels. In combination with theoretical, biophysical and pharmacological studies, we learn about the basic mechanism of ion permeation and channel gating at the molecular level. One of the objectives is to understand the unique permeation properties of BK channels, which allow K+ to permeate 20-fold better in respect to other potassium channels that share very similar amino-acid pore sequences.
We are developing a collaborative research program focused on cardiac arrhythmias and fibrillation. Activation and inactivation of voltage-dependent calcium channels play a key role in the stability of cardiac rhythm, through their effects on action-potential properties. The goal is to is to control cardiac excitability by fine tuning the the activity of cardiac L-type calcium channels, using their own modulatory subunits. The project is intended as the foundation for a larger gene therapy-related project.
Dr. Olcese holds the rank of Professor in the Departments of Physiology and of Anesthesiology & Perioperative Medicine, and is Director of Basic Science Research Program Administration. He is affiliated with the UCLA Brain Research Institute, the graduate program in Molecular, Cellular, and Integrative Physiology, the graduate program in Neuroscience, and the Cardiovascular Research Laboratory.
Savalli N*, Pantazis A, Sigg D, Weiss JN, Neely A, Olcese R. The α2δ-1 subunit remodels CaV1.2 voltage sensors and allows Ca2+ influx at physiological membrane potentials. J Gen Physiol. 2016;148(2):147-59 PMCID: PMC4969795. (Recommended by "Faculty of 1000 Prime", *Awarded 2017 Cranefield Award for this work)
Voltage-activated ion channels, are largely responsible for cell electrical excitability. They “sense” the electric potential across the plasmamembrane using specialized Voltage Sensing Domains that compel the pore to open it. By optically tracking the movement of a human calcium channel using a technique called Voltage Clamp fluorometry, the Olcese laboratory revealed the mechanism by which one accessory proteins (called a2d-1) increases the sensitivity of the channel to membrane depolarizations, thus allowing the channel to operate at physiological potentials. This work has been highlighted by a Commentary in the same issue of the Journal of General Physiology, and by Dr. A. C. Dolphin who recommended the work as being of special significance in the Faculty of 1000 Prime.
Pantazis A, Olcese R. Biophysics of BK Channel Gating. Int Rev Neurobiol. 2016; 128:1-49 PMID: 27238260.
BK channels are a central focus of the Olcese laboratory. They are universal regulators of cell excitability, given their exceptional unitary conductance selective for K+, joint activation mechanism by membrane depolarization and intracellular [Ca2+] elevation, and broad expression pattern. In this chapter, we discuss the structural basis and operational principles of their activation, or gating, by membrane potential and calcium. We also discuss how the two activation mechanisms interact to culminate in channel opening. As members of the voltage-gated potassium channel superfamily, BK channels are discussed in the context of archetypal family members, in terms of similarities that help us understand their function, but also seminal structural and biophysical differences that confer unique functional properties.
Karagueuzian HS, Pezhouman A, Angelini M, Olcese R. Enhanced Late Na and Ca Currents as Effective Antiarrhythmic Drug Targets. Front Pharmacol. 2017; 6;8:36. PMCID: PMC5292429.
Pathological rise of either or both of INa-Land late ICa,Lare demonstrated to promote cellular early afterdepolarizations (EADs) and EAD-mediated triggered activity that can initiate VT/VF in remodeled hearts. The selective inhibition of the INa-L without affecting peak with the highly specific prototype drug GS-967 suppresses EAD-mediated VT/VFs. As in the case of INa-L, selective inhibition of the late ICa,Lwithout affecting its peak with the prototype drug, roscovitine suppressed oxidative EAD-mediated VT/VF. These findings indicate that specific blockers of the late inward currents without affecting their peaks (gating modifiers), offer a new and effective AAD class action i.e., "Class VI." The development of safe drugs with selective Class VI actions provides a rational and effective approach to treat VT/VF particularly in cardiac conditions associated with enhanced CaMKII activity such as heart failure.
Zhang J, Li M, Zhang Z, Zhu R, Olcese R, Stefani E, Toro L. The mitochondrial BK(Ca) channel cardiac interactome reveals BK(Ca) association with the mitochondrial import receptor subunit Tom22, and the adenine nucleotide translocator. Mitochondrion 2017; 33:84-101. PMCID: PMC5332438.
In this collaborative effort, a directed proteomic approach discovered the novel interaction of the large-conductance Ca- and voltage-activated BKCa channel with Tom22, a component of the mitochondrion outer membrane import system, and the adenine nucleotide translocator (ANT). The results from this work demonstrate provide a molecular framework to understand the intricacies of mitoBKCa mitochondrial import mechanism and function.