Intracellular β1-Adrenergic Receptors and Organic Cation Transporter 3 Mediate Phospholamban Phosphorylation to Enhance Cardiac Contractility

Rationale: β 1 -adrenoceptors (β 1 ARs) exist at intracellular membranes and Organic Cation Transporter 3 (OCT3) mediates norepinephrine entry into cardiomyocytes. However, the functional role of intracellular β 1 AR in cardiac contractility remains to be elucidated. Objective: Test localization and function of intracellular β 1 AR on cardiac contractility. Methods and Results: Membrane fractionation, super-resolution imaging, proximity ligation, co-immunoprecipitation and single-molecule pulldown demonstrated a pool of β 1 ARs in mouse hearts that was associated with sarco/endoplasmic reticulum Ca 2+ -ATPase at the sarcoplasmic reticulum (SR). Local protein kinase A (PKA) activation was measured using a PKA biosensor targeted at either the plasma membrane (PM) or SR. Compared to wild type (WT), myocytes lacking OCT3 (OCT3KO) responded identically to the membrane-permeant βAR agonist isoproterenol in PKA activation at both PM and SR. The same was true at the PM for membrane-impermeant norepinephrine, but the SR response to norepinephrine was suppressed in OCT3KO myocytes. This differential effect was recapitulated in phosphorylation of the SR-pump regulator phospholamban. Similarly, OCT3KO selectively suppressed calcium transients and contraction responses to norepinephrine, but not isoproterenol. Furthermore, sotalol, a membrane-impermeant βAR-blocker suppressed isoproterenol-induced PKA activation at the PM, but permitted PKA activation at the SR, phospholamban phosphorylation and contractility. Moreover, pretreatment with sotatol in OCT3KO myocytes prevented norepinephrine induced PKA activation at both PM and the SR and contractility. Conclusions: Functional β 1 ARs exists at the SR and is critical for PKA-mediated phosphorylation of phospholamban and cardiac contractility upon catecholamine stimulation. Activation of these intracellular β 1 ARs requires catecholamine transport via OCT3.

Download Full-text

Organic cation transporter 3 ist im hepatischen Kupfermetabolismus involviert

Zeitschrift für Gastroenterologie ◽

10.1055/s-0038-1668784 ◽

2018 ◽

Vol 56 (08) ◽

pp. e250-e250

Author(s):

S Guttmann ◽

S Reinartz Groba ◽

C Niemietz ◽

V Sandfort ◽

A Zibert ◽

...

Keyword(s):

Organic Cation ◽

Organic Cation Transporter ◽

Organic Cation Transporter 3

Download Full-text

Super-Resolution Imaging and Single Molecule Tracking of the Nuclear Protein Emerin

Biophysical Journal ◽

10.1016/j.bpj.2013.11.1186 ◽

2014 ◽

Vol 106 (2) ◽

pp. 201a

Author(s):

Anthony M. Fernandez ◽

Fabien F. Pinaud

Keyword(s):

Single Molecule ◽

Nuclear Protein ◽

Super Resolution ◽

Single Molecule Tracking ◽

Resolution Imaging

Download Full-text

Prenatal metformin exposure or organic cation transporter 3 knock-out curbs social interaction preference in male mice

Pharmacological Research ◽

10.1016/j.phrs.2018.11.013 ◽

2019 ◽

Vol 140 ◽

pp. 21-32 ◽

Cited By ~ 2

Author(s):

Valentina R. Garbarino ◽

Taylor A. Santos ◽

Anastassia R. Nelson ◽

Wynne Q. Zhang ◽

Corey M. Smolik ◽

...

Keyword(s):

Social Interaction ◽

Organic Cation ◽

Organic Cation Transporter ◽

Male Mice ◽

Knock Out ◽

Organic Cation Transporter 3

Download Full-text

Regulation Mechanisms of Expression and Function of Organic Cation Transporter 1

Frontiers in Pharmacology ◽

10.3389/fphar.2020.607613 ◽

2021 ◽

Vol 11 ◽

Author(s):

Giuliano Ciarimboli

Keyword(s):

Organic Cation ◽

Basolateral Membrane ◽

Organic Cation Transporter ◽

Organic Cations ◽

Regulatory Processes ◽

Organic Cation Transporter 1 ◽

Exogenous Origin ◽

And Function

The organic cation transporter 1 (OCT1) belongs together with OCT2 and OCT3 to the solute carrier family 22 (SLC22). OCTs are involved in the movement of organic cations through the plasma membrane. In humans, OCT1 is mainly expressed in the sinusoidal membrane of hepatocytes, while in rodents, OCT1 is strongly represented also in the basolateral membrane of renal proximal tubule cells. Considering that organic cations of endogenous origin are important neurotransmitters and that those of exogenous origin are important drugs, these transporters have significant physiological and pharmacological implications. Because of the high expression of OCTs in excretory organs, their activity has the potential to significantly impact not only local but also systemic concentration of their substrates. Even though many aspects governing OCT function, interaction with substrates, and pharmacological role have been extensively investigated, less is known about regulation of OCTs. Possible mechanisms of regulation include genetic and epigenetic modifications, rapid regulation processes induced by kinases, regulation caused by protein–protein interaction, and long-term regulation induced by specific metabolic and pathological situations. In this mini-review, the known regulatory processes of OCT1 expression and function obtained from in vitro and in vivo studies are summarized. Further research should be addressed to integrate this knowledge to known aspects of OCT1 physiology and pharmacology.

Download Full-text

Comparing lifeact and phalloidin for super-resolution imaging of actin in fixed cells

PLoS ONE ◽

10.1371/journal.pone.0246138 ◽

2021 ◽

Vol 16 (1) ◽

pp. e0246138

Author(s):

Hanieh Mazloom-Farsibaf ◽

Farzin Farzam ◽

Mohamadreza Fazel ◽

Michael J. Wester ◽

Marjolein B. M. Meddens ◽

...

Keyword(s):

Single Molecule ◽

Cell Biology ◽

Cell Movement ◽

Super Resolution ◽

Actin Binding ◽

Thin Filaments ◽

Reversible Binding ◽

Multiple Regions ◽

Resolution Imaging ◽

Division Cell

Visualizing actin filaments in fixed cells is of great interest for a variety of topics in cell biology such as cell division, cell movement, and cell signaling. We investigated the possibility of replacing phalloidin, the standard reagent for super-resolution imaging of F-actin in fixed cells, with the actin binding peptide ‘lifeact’. We compared the labels for use in single molecule based super-resolution microscopy, where AlexaFluor 647 labeled phalloidin was used in a dSTORM modality and Atto 655 labeled lifeact was used in a single molecule imaging, reversible binding modality. We found that imaging with lifeact had a comparable resolution in reconstructed images and provided several advantages over phalloidin including lower costs, the ability to image multiple regions of interest on a coverslip without degradation, simplified sequential super-resolution imaging, and more continuous labeling of thin filaments.

Download Full-text

The architecture and operating mechanism of a cnidarian stinging organelle

10.21203/rs.3.rs-888152/v1 ◽

2021 ◽

Author(s):

Matthew Gibson ◽

Ahmet Karabulut ◽

Melainia McClain ◽

Boris Rubinstein ◽

Sean McKinney

Keyword(s):

Structural Complexity ◽

Super Resolution ◽

Operating Mechanism ◽

Sea Anemones ◽

Form And Function ◽

3D Electron Microscopy ◽

Genetic Perturbations ◽

Resolution Imaging ◽

The Operating Mechanism ◽

And Function

Abstract The stingers of jellyfish, sea anemones and other cnidarians, known as nematocysts, are remarkable cellular weapons used for both predation and defense1. Nematocysts are specialized organelles which consist of a pressurized capsule containing a coiled harpoon-like thread2. These structures are in turn built within specialized cells known as nematocytes3. When triggered4, the capsule explosively discharges, ejecting the coiled thread which punctures the target and rapidly elongates by turning inside out in a process called eversion5,6. Due to the structural complexity of the thread and the extreme speed of discharge, the precise mechanics of nematocyst firing have remained elusive7. Here, using a combination of live and super-resolution imaging, 3D electron microscopy and genetic perturbations, we define the step-by-step sequence of nematocyst operation in the model sea anemone Nematostella vectensis. This analysis reveals the complex biomechanical transformations underpinning the operating mechanism of nematocysts, one of the nature’s most exquisite biological micro-machines. Further, this study will provide insight into the form and function of related cnidarian organelles and serve as a template for the design of bioinspired microdevices.

Download Full-text