Large Acid-Evoked Currents, Mediated by ASIC1a, Accompany Differentiation in Human Dopaminergic Neurons.

Acid-sensing ion channels (ASICs) are proton-gated Na+ channels. They contribute to synaptic transmission, neuronal differentiation and neurodegeneration. ASICs have been mainly characterized in neurons from mice or rats and our knowledge of their properties in human neurons is scarce. Here, we functionally characterized ASICs in differentiating LUHMES cells, a human mesencephalic cell line with characteristics of dopaminergic neurons. We find that LUHMES cells express functional ASICs, predominantly homomeric ASIC1a. Expression starts early during differentiation with a striking surge in current amplitude at days 4-6 of differentiation, a time point where-based on published data-LUHMES cells start expressing synaptic markers. Peak ASIC expression therefore coincides with a critical period of LUHMES cell differentiation. It was associated with increased excitability, but not paralleled by an increase in ASIC1 mRNA or protein. In differentiating as well as in terminally differentiated LUHMES cells, ASIC activation by slight acidification elicited large currents, action potentials and a rise in cytosolic Ca2+. Applying the ASIC pore blocker diminazene during differentiation reduced the length of neurites, consistent with the hypothesis that ASICs play a critical role in LUHMES cell differentiation. In summary, our study establishes LUHMES cells as a valuable model to study the role of ASICs for neuronal differentiation and potentially also cell death in a human cell line.

Glioblastoma cancer stem cell lines express functional acid sensing ion channels ASIC1a and ASIC3.

Acidic microenvironment is commonly observed in tumour tissues, including glioblastoma (GBM), the most aggressive and lethal brain tumour in adults. Acid sensing ion channels (ASICs) are neuronal voltage-insensitive sodium channels, which are sensors of extracellular protons. Here we studied and functionally characterized ASICs in two primary glioblastoma stem cell lines as cell culture models. We detected transcripts of the ACCN2 and ACCN3 genes, coding for ASIC1 and ASIC3, respectively, but not transcripts of ACCN1 (coding for ASIC2). Available microarray data confirmed that ACCN1 is downregulated in glioma. Western blotting confirmed expression of ASIC1 and ASIC3, the most proton-sensitive ASICs, in both GBM cell lines. We characterized ASICs functionally using whole-cell patch clamp and detected different types of acid-sensitive currents. Some of these currents had kinetics typical for ASICs and were sensitive to specific toxin inhibitors of ASIC1a or ASIC3, demonstrating that the GBM cell lines express functional ASIC1a and ASIC3 that may enable GBM cells to sensitively detect extracellular pH in a tumour tissue. Microarray data revealed that expression of ACCN2 and ACCN3 is associated with improved survival of patients suffering from gliomas, suggesting that preserved susceptibility to extracellular pH may impair tumour growth.

Ultrathin Coatings with Change in Reactivity over Time Enable Functional In Vitro Networks Of Insect Neurons.

It's just not cricket! A novel coating system that enables covalent attachment of biomolecules in a nonfouling environment without use of additional chemical crosslinkers is presented. Concanavalin A is patterned on the coatings to direct cell adhesion and growth of neurons from the cricket Gryllus bimaculatus and generate functional, patterned in vitro insect neuronal networks for the first time.

Surface grafting of PEO-based star-shaped molecules for bioanalytical and biomedical applications.

This article reviews surface grafting of star-shaped PEO. The use of star-shaped polymers is compared to linear PEO chains regarding the layer preparation and the ability of the resulting surfaces to resist protein adsorption. We then focus on the use of end-functionalized, star-shaped, PEO-based prepolymers that are able to form covalent crosslinks and functional polymer networks on the substrate. Examples are given for specific protein adsorption as well as for cell adhesion on such layers by covalent embedding of biofunctional molecules. The possibility of coating biomedically relevant polymer substrates in three-dimensional geometries is discussed and examples are shown for poly(ethylene terephthalate) monofilament constructs.

Microcontact printing of proteins for neuronal cell guidance.

The growth of neurons into networks of controlled geometry is of great interest in the field of cell-based biosensors, neuroelectronic circuits, neurological implants, pharmaceutical testing as well as fundamental biological questions about neuronal interactions. The precise control of the network architecture can be achieved by defined engineering of the surface material properties: this process is called neuronal cell patterning. Different techniques can be used to produce such surface patterns. We have chosen microcontact printing (µCP), because it is a comparatively simple and universal method for patterning biomolecules.