The ß-cell primary cilium is an autonomous Ca2+ compartment for paracrine GABA signaling.

The primary cilium is an organelle present in most adult mammalian cells that is considered as an antenna for sensing the local microenvironment. Here, we use intact mouse pancreatic islets of Langerhans to investigate signaling properties of the primary cilium in insulin-secreting ß-cells. We find that GABAB1 receptors are strongly enriched at the base of the cilium, but are mobilized to more distal locations upon agonist binding. Using cilia-targeted Ca2+ indicators, we find that activation of GABAB1 receptors induces selective Ca2+ influx into primary cilia through a mechanism that requires voltage-dependent Ca2+ channel activation. Islet ß-cells utilize cytosolic Ca2+ increases as the main trigger for insulin secretion, yet we find that increases in cytosolic Ca2+ fail to propagate into the cilium, and that this isolation is largely due to enhanced Ca2+ extrusion in the cilium. Our work reveals local GABA action on primary cilia that involves Ca2+ influx and depends on restricted Ca2+ diffusion between the cilium and cytosol.

Quantification of Secretory Granule Exocytosis by TIRF Imaging and Capacitance Measurements.

Hormones and neurotransmitters are released from (neuro)endocrine cells by regulated exocytosis of secretory granules. During exocytosis, the granule membrane fuses with the plasma membrane, which allows release of the stored content into the bloodstream or the surrounding tissue. Here, we give a detailed description of two complementary methods to observe and quantify exocytosis in single cells: high-resolution TIRF microscopy and patch-clamp capacitance recordings. Precise stimulation of exocytosis is achieved by local pressure application or voltage-clamp depolarizations. While the chapter is focused on insulin-secreting cells as an accessible and disease-relevant model system, the methodology is applicable to a wide variety of secretory cells including chromaffin and PC12 cells.

Activation of BK Channel Contributes to PL-Induced Mesenchymal Stem Cell Migration.

Due to their capacity to proliferate, migrate, and differentiate, mesenchymal stem cells (MSCs) are considered to be good candidates for regenerative medicine applications. The mechanisms underlying proliferation and differentiation of MSCs have been studied. However, much less is known about the mechanisms regulating the migration of MSCs. Platelet lysate (PL), a supplement used to promote cell expansion, has been shown to promote MSCs migration; however, the underlying mechanism are unknown. Here, by using adipose-derived rat MSCs (rMSCs) and the scratch assay in the absence and presence of various BK channels modulators, we evaluated the role of BK channels in mediating the PL-stimulated migration of rMSCs. We found that 5% PL increased rMSCs migration, and this effect was blocked by the addition of the BK channel selective antagonist Iberiotoxin (IBTX). In the absence of PL, the BK channel agonist NS1619, stimulated rMSCs migration to similar level as 5% PL. Addition of both NS1619 and 5% PL resulted in an increase in rMSCs migration, that was higher than when either one was added individually. From whole-cell recordings, it was found that the addition of 5% PL increased the magnitude of BK current density. By using Western blot and flow cytometry, it was found that PL did not affect the expression of BK channels. Together, our results indicate that as shown in other cell types, activation of BK channels by themselves also promote rMSC migration, and show that activation of BK channels contribute to the observed PL-induced increase in migration of rMSC.

Transient Receptor Potential Channels in Microglia: Roles in Physiology and Disease.

Microglia modulate the nervous system cellular environment and induce neuroprotective and neurotoxic effects. Various molecules are involved in these processes, including families of ion channels expressed in microglial cells, such as transient receptor potential (TRP) channels. TRP channels comprise a family of non-selective cation channels that can be activated by mechanical, thermal, and chemical stimuli, and which contribute to the regulation of intracellular calcium concentrations. TRP channels have been shown to be involved in cellular processes such as osmotic regulation, cytokine production, proliferation, activation, cell death, and oxidative stress responses. Given the significance of these processes in microglial activity, studies of TRP channels in microglia have focused on determining their roles in both neuroprotective and neurotoxic processes. TRP channel activity has been proposed to play an important function in neurodegenerative diseases, ischemia, inflammatory responses, and neuropathic pain. Modulation of TRP channel activity may thus be considered as a potential therapeutic strategy for the treatment of various diseases associated with alterations of the central nervous system (CNS). In this review, we describe the expression of different subfamilies of TRP channels in microglia, focusing on their physiological and pathophysiological roles, and consider their potential use as therapeutic targets in CNS diseases.

Pilot study evaluating the safety of a combined central venous catheter and inferior vena cava filter in critically ill patients at high risk of pulmonary embolism.

The objectives of this pilot trial were to assess the safety of a new device for pulmonary embolism (PE) prophylaxis. The device, the Angel Catheter, was placed in eight patients who were in the intensive care unit and were at high risk of PE. The device was inserted at the bedside without fluoroscopic guidance via a femoral venous approach. All eight devices were inserted and subsequently retrieved without complications (follow-up, 33-36 d). One filter trapped a large clot.