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Contribution of electromechanical coupling between Kv and Ca v1.2 channels to coronary dysfunction in obesity.

ABSTRACT: Previous investigations indicate that diminished functional expression of voltage-dependent K(+) (KV) channels impairs control of coronary blood flow in obesity/metabolic syndrome. The goal of this investigation was to test the hypothesis that KV channels are electromechanically coupled to CaV1.2 channels and that coronary microvascular dysfunction in obesity is related to subsequent increases in CaV1.2 channel activity. Initial studies revealed that inhibition of KV channels with 4-aminopyridine (4AP, 0.3 mM) increased intracellular [Ca(2+)], contracted isolated coronary arterioles and decreased coronary reactive hyperemia. These effects were reversed by blockade of CaV1.2 channels. Further studies in chronically instrumented Ossabaw swine showed that inhibition of CaV1.2 channels with nifedipine (10 ?g/kg, iv) had no effect on coronary blood flow at rest or during exercise in lean swine. However, inhibition of CaV1.2 channels significantly increased coronary blood flow, conductance, and the balance between coronary flow and metabolism in obese swine (P < 0.05). These changes were associated with a ~50 % increase in inward CaV1.2 current and elevations in expression of the pore-forming subunit (?1c) of CaV1.2 channels in coronary smooth muscle cells from obese swine. Taken together, these findings indicate that electromechanical coupling between KV and CaV1.2 channels is involved in the regulation of coronary vasomotor tone and that increases in CaV1.2 channel activity contribute to coronary microvascular dysfunction in the setting of obesity.

SUBMITTER: Berwick ZC

PROVIDER: S-EPMC3818702 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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Contribution of electromechanical coupling between Kv and Ca v1.2 channels to coronary dysfunction in obesity.

Berwick Zachary C ZC Dick Gregory M GM O'Leary Heather A HA Bender Shawn B SB Goodwill Adam G AG Moberly Steven P SP Owen Meredith Kohr MK Miller Steven J SJ Obukhov Alexander G AG Tune Johnathan D JD

Basic research in cardiology 20130716 5

Previous investigations indicate that diminished functional expression of voltage-dependent K(+) (KV) channels impairs control of coronary blood flow in obesity/metabolic syndrome. The goal of this investigation was to test the hypothesis that KV channels are electromechanically coupled to CaV1.2 channels and that coronary microvascular dysfunction in obesity is related to subsequent increases in CaV1.2 channel activity. Initial studies revealed that inhibition of KV channels with 4-aminopyridin ...[more]

PMID: 23856709

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Project description:Voltage-dependent potassium channels (Kv) are extremely sensitive to membrane voltage and play a crucial role in membrane repolarization during action potentials. Kv channels undergo voltage-dependent transitions between closed states before opening. Despite all we have learned using electrophysiological methods and structural studies, we still lack a detailed picture of the energetics of the activation process. We show here that even a single mutation can drastically modify the temperature response of the Shaker Kv channel. Using rapid cell membrane temperature steps (Tsteps), we explored the effects of temperature on the ILT mutant (V369I, I372L, and S376T) and the I384N mutant. The ILT mutant produces a significant separation between the transitions of the voltage sensor domain (VSD) activation and the I384N uncouples its movement from the opening of the domain (PD). ILT and I384N respond to temperature in drastically different ways. In ILT, temperature facilitates the opening of the channel akin to a "hot" receptor, reflecting the temperature dependence of the voltage sensor's last transition and facilitating VSD to PD coupling (electromechanical coupling). In I384N, temperature stabilizes the channel closed configuration analogous to a "cold" receptor. Since I384N drastically uncouples the VSD from the pore opening, we reveal the intrinsic temperature dependence of the PD itself. Here, we propose that the electromechanical coupling has either a "loose" or "tight" conformation. In the loose conformation, the movement of the VSD is necessary but not sufficient to efficiently propagate the electromechanical energy to the S6 gate. In the tight conformation the VSD activation is more effectively translated into the opening of the PD. This conformational switch can be tuned by temperature and modifications of the S4 and S4-S5 linker. Our results show that we can modulate the temperature dependence of Kv channels by affecting its electromechanical coupling.

Project description:Voltage-gated ion channels play a central role in the generation of action potentials in the nervous system. They are selective for one type of ion - sodium, calcium, or potassium. Voltage-gated ion channels are composed of a central pore that allows ions to pass through the membrane and four peripheral voltage sensing domains that respond to changes in the membrane potential. Upon depolarization, voltage sensors in voltage-gated potassium channels (Kv) undergo conformational changes driven by positive charges in the S4 segment and aided by pairwise electrostatic interactions with the surrounding voltage sensor. Structure-function relations of Kv channels have been investigated in detail, and the resulting models on the movement of the voltage sensors now converge to a consensus; the S4 segment undergoes a combined movement of rotation, tilt, and vertical displacement in order to bring 3-4e(+) each through the electric field focused in this region. Nevertheless, the mechanism by which the voltage sensor movement leads to pore opening, the electromechanical coupling, is still not fully understood. Thus, recently, electromechanical coupling in different Kv channels has been investigated with a multitude of techniques including electrophysiology, 3D crystal structures, fluorescence spectroscopy, and molecular dynamics simulations. Evidently, the S4-S5 linker, the covalent link between the voltage sensor and pore, plays a crucial role. The linker transfers the energy from the voltage sensor movement to the pore domain via an interaction with the S6 C-termini, which are pulled open during gating. In addition, other contact regions have been proposed. This review aims to provide (i) an in-depth comparison of the molecular mechanisms of electromechanical coupling in different Kv channels; (ii) insight as to how the voltage sensor and pore domain influence one another; and (iii) theoretical predictions on the movement of the cytosolic face of the Kv channels during gating.

Project description:The recently determined crystal structure of a chimeric Kv1.2-Kv2.1 Kv channel at 2.4 A resolution motivated this molecular-dynamics simulation study of the chimeric channel and its mutants embedded in a DPPC membrane. For the channel protein, we used two types of C-terminus: E+ and Eo. E+ contains, and Eo lacks, the EGEE residue quartet located distal to the S6 helix. For both E+ and Eo, the following trend was observed: When S4 helices were restrained at the same position as in the x-ray structure (S4high), the S6 gate remained open for 12 ns. The results were similar when the S4 helices were pulled downward 7 A (S4low). However, S4middle (or S4low) facilitated the S6 gate-narrowing for the following mutated channels (shown in order of increasing effect): 1), E395W; 2), E395W-F401A-F402A; and 3), E395W-F401A-F402A-V478W. The amino acid numbering system is that used for the Shaker channel. Even though all four subunits were set at S4low, S6 gate-narrowing was often brought about by movements of only two opposing S6 helices toward the central axis of the pore, resulting in a twofold symmetry-like structure. A free-energy profile analysis over the ion conduction pathway shows that the two opposing S6 helices whose peptide backbones are approximately 10.4 A distant from each other lead to an energetic barrier of approximately 25 kJ/mol. S6 movement was coupled with translocation of the S4-S5 linker toward the central axis of the same subunit, and the coupling was mediated by salt bridges formed between the inner (intracellular side) end of S4 and that of S6. Simulations in which S4 of only one subunit was pulled down to S4low showed that a weak intersubunit coordination is present for S5 movement, whereas the coupling between the S4-S5 linker and S6 is largely an intrasubunit one. In general, whereas subunit-based behavior appears to be dominant and to permit heteromeric conformations of the pore domain, direct intersubunit coupling of S5 or S6 is weak. Therefore, the "concerted transition" of the pore domain that has been predicted based on electrophysiological analyses is likely to be mediated mainly by the dual effects of S4 and the S4-S5 linker; these segments of one subunit can interact with both S5 of the same subunit and that of the adjacent subunit.

Dataset Information

Contribution of electromechanical coupling between Kv and Ca v1.2 channels to coronary dysfunction in obesity.

Publications

Contribution of electromechanical coupling between Kv and Ca v1.2 channels to coronary dysfunction in obesity.

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