This symposium is organized by the Young Physiologists for the three ‚Best Talks’ Awardees of the 7th Young Physiologists Symposium in Nuremberg in March this year. They will give their selected talks following introductions by well-established experts from their field of research.
Thyroid hormones in ocular angiogenesis and inflammation – implications for retinal degeneration (#473)
Y. Henning1, S. Plöhn2, A. Eckstein2, U. Berchner-Pfannschmidt2, J. Fandrey1
1 University of Duisburg-Essen, University Hospital Essen, Institute of Physiology, Essen, Germany
Many people suffer from retinal degenerative diseases with different levels of vision loss up to complete blindness. Currently, thyroid hormones (TH), in particular triiodothyronine (T3) and its precursor thyroxine (T4), gain broader attention as crucial factors for retinal development and photoreceptor function. For instance, alterations of the TH status influence color vision, and excess TH signaling was previously linked to degeneration of cone photoreceptors. However, the molecular mechanisms causative for these degenerative processes are not clearly identified thus far. Retinal diseases are often associated with neovascularization and inflammatory processes, such as in age-related macular degeneration, the most common cause of blindness in elderly people, where central vision is lost. Since TH have proangiogenic and proinflammatory effects, we are currently investigating a potential link between TH signaling, neovascularization and inflammation in the retina. In the presented project, we are focusing on the role of the retinal pigment epithelium (RPE), a monolayer of pigmented cells located adjacent to the retina, in photoreceptor degeneration, because the RPE is crucial for maintaining visual function by regulating photoreceptor homeostasis. To obtain the first insights into TH signaling in the RPE, we treated an immortalized RPE cell line (ARPE-19) with T3 and observed proangiogenic and proinflammatory T3 effects. These effects are likely to be driven by hypoxia-inducible factor 1 (HIF-1), a key regulator of proangiogenic processes under hypoxic conditions, because we found that T3 treatment upregulated HIF-1α, the α-subunit of HIF-1. Moreover, preliminary data on a mouse model with endogenous hyperthyroidism support our findings on TH-induced proangiogenic effects.
Resurgent-like sodium currents induced by insecticide in the absence of pore-blocker (#472)
S. Thull1, C. Neacsu2, A. O. O'Reilly3, T. Huth2, J. Meents1, A. Lampert1
1 RWTH Aachen University, Institute of Physiology, Aachen, Germany
Voltage-gated sodium channels are responsible not only for the fast upstroke of the action potential, but they also modify cellular excitability via persistent and resurgent currents. We are beginning to understand the molecular basis of the latter: it is commonly accepted that an open channel blocker is necessary to interfere with the channel’s inactivation process to produce resurgent currents. Here, we applied the classical type-II pyrethroid deltamethrin on human cardiac Nav1.5 expressed in HEK cells during whole-cell patch-clamp experiments. Surprisingly, we observed resurgent-like currents at very negative potentials in the absence of any open-channel blocker. Deltamethrin is predicted to bind with and stabilize domain II of the channel in the activated position to inhibit sodium channel deactivation. In contrast, activation of domain IV is associated with binding of the inactivation particle that terminates ion flow. Our experiments suggest that in deltamethrin-modified channels, resurgent-like currents are generated because the movement of domain IV and the resulting unbinding of the inactivation particle occur quicker than deactivation via movement of domain II. Notably, when activation of domain IV was inhibited by co-application of the sea anemone toxin ATx-II, these resurgent-like currents were eliminated although persistent currents were still observed. This indicates that the channel is permeable even if domain IV does not activate and generation of resurgent-like currents requires domain IV movement. By illuminating additional conducting sodium channel gating states and giving evidence for an alternative way of resurgent current induction in sodium channels, our findings shed new light on the complex sodium channel gating.
Environmental pH and mechanical stress orchestrate pancreatic stellate cell activation (#471)
Z. Pethö1, B. Fels1, 2, A. Schwab1
1 University of Münster, Institute of Physiology II, Münster, Germany
Pancreatic stellate cells (PSCs) are stromal cells of the pancreas that can become activated in various diseases such as pancreatic cancer. Pancreatic cancer is coupled to changes in the extracellular environment of the stroma, including mechanical stiffness and interstitial pH. The tumorous transformation of the pancreatic ductal cells leads to impaired ductal HCO3- secretion, which in turn leads to a relative alkalinization of the interstitial pH of the pancreas. In this study we aim to elucidate how changes in extracellular pH (pHe) and mechanical stress affect the intracellular pH (pHi) homeostasis and activation of pancreatic stellate cells.
We isolated PSCs from healthy adult wild-type C57/BL6 mice. Following isolation, we cultured PSCs for up to 120 hours in media buffered to different pHe on a stiff substrate or on polyacrylamide gels of physiological stiffness. Cellular viability, activation and proliferation were assessed using immunocytochemistry and flow cytometry. To investigate the functional expression of potential pH sensors, regulators and pH-regulated transporters of PSCs, we performed RT-qPCR, Western blot, immunofluorescence and pHi measurements.
Alkalization of the extracellular milieu substantially facilitated PSC activation as evidenced by an increase in cell size and α-SMA positivity, especially on a rigid substrate. Similarly, cell proliferation, cell cycle progression and nuclear Yap localization are also facilitated in an alkaline pHe environment. Multiple pH sensors and regulators are highly expressed in activated PSCs cultured in pHe 7.4 compared to freshly isolated cells and cells cultured in pHe 6.6. In addition, NHE1 activity is enhanced in activated PSCs at pHe 7.4 as compared to inactive PSCs at pHe 6.6. When PSCs are cultured in an acidic pHe environment, the pHi acidifies to pHi ≤ 7.0. In summary, alkaline pHe is a potent activator of PSCs, that may enable cell cycle progression through Yap/Taz and HIF1a mediated pathways.