Stroke is a leading cause of death and long-term disabilities. Despite decades of extensive efforts in search of brain injury mechanisms and therapeutic interventions, pharmacological treatment is limited to the use of thrombolytic agent tissue plasminogen activator, which has limited therapeutic time window and potential side effect of intracranial hemorrhage. Over the past few years, endovascular thrombectomy with stent-retriever devices combined with advanced imaging modalities has transformed the standard of stroke care, offering an opportunity to improve the outcome in selected patients as late as 24 h after the onset of stroke. This mini-review summarizes the advancement in the treatment of ischemic stroke, from thrombolysis to thrombectomy and remaining challenges in the field.

High-glucose (HG) levels and hyperglycemia associated with diabetes are known to cause neuronal damage. The detailed molecular mechanisms, however, remain to be elucidated. Here, we investigated the role of transient receptor potential melastatin 7 (TRPM7) channels in HG-mediated endoplasmic reticulum stress (ERS) and injury of NS20Y neuronal cells. The cells were incubated in the absence or presence of HG for 48 h. We found that mRNA and protein levels of TRPM7 and of ERS-associated proteins such as C/EBP homologous protein (CHOP), 78 KDa glucose-regulated protein (GRP78), and inducible nitric oxide synthase (iNOS) increased in HG-treated cells, along with significantly increased TRPM7-associated currents in these cells. Similar results were obtained in cerebral cortical tissue from an insulin-deficiency model of diabetic mice. Moreover, HG treatment of cells activated ERS-associated proapoptotic caspase activity and induced cellular injury. Interestingly, a NOS inhibitor, L-NAME, suppressed the HG-induced increase of TRPM7 expression and cellular injury. siRNA-mediated TRPM7 knockdown or chemical inhibition of TRPM7 activity also suppressed HG-induced ERS and decreased cleaved caspase12/caspase3 levels and cell injury. Of note, TRPM7 overexpression increased ERS and cell injury independently of its kinase activity. Taken together, our findings suggest that TRPM7 channel activities play a key role in HG-associated ERS and cytotoxicity through an apoptosis-inducing signaling cascade involving HG, iNOS, TRPM7, other ERS proteins, and caspases.

Background: Extracellular acidosis is a prominent feature of multiple pathological conditions, correlating with pain sensation. Acid-sensing ion channels (ASICs), a family of proton-gated cation channels, are distributed throughout the central and peripheral nervous systems. Activation of ASICs, particularly ASIC3 and ASIC1a channels, by acidic pH and the resultant depolarization of nociceptive primary sensory neurons, participates in nociception. Agents that inhibit the activation of ASICs are thus expected to be analgesic. Here, we studied the effect of local anesthetic tetracaine on ASIC currents. Results: Tetracaine inhibited the peak ASIC3 current in a concentration-dependent manner with an IC₅₀ of 9.96 ± 1.88 mM. The degree of inhibition by tetracaine was dependent on the extracellular pH but independent of the membrane potential. Furthermore, 3 mM tetracaine also inhibited 29.83% of the sustained ASIC3 current. In addition to ASIC3, tetracaine inhibited the ASIC1a and ASIC1β currents. The inhibition of the ASIC1a current was influenced by the frequency of channel activation. In contrast to ASIC3, ASIC1a, and ASIC1β currents, ASIC2a current was not inhibited by tetracaine. In cultured mouse dorsal root ganglion neurons, 1–3 mM tetracaine inhibited both the transient and sustained ASIC currents. At pH4.5, 3 mM tetracaine reduced the peak ASIC current to 60.06 ± 4.51%, and the sustained current to 48.24 ± 7.02% of the control values in dorsal root ganglion neurons. In contrast to ASICs, voltage-gated sodium channels were inhibited by acid, with 55.15% inhibition at pH6.0 and complete inhibition at pH5.0. Conclusions: These findings disclose a potential new mechanism underlying the analgesic effects of local anesthetics, particularly in acidic conditions where their primary target (i.e. voltage-gated Na⁺ channel) has been suppressed by protons.

Acid-sensing ion channels (ASICs) are proton-gated cation channels that mediate acid-induced responses in neurons. ASICs are important for mechanosensation, learning and memory, fear, pain, and neuronal injury. ASIC2a is widely expressed in the nervous system and modulates ASIC channel trafficking and activity in both central and peripheral systems. Here, to better understand mechanisms regulating ASIC2a, we searched for potential protein motifs that regulate ASIC2a trafficking. We identified a LLDLL sequence in the C-terminal juxtamembrane region of ASIC2a. Deleting or mutating the LLDLL sequence increased total expression and surface levels of ASIC2a in CHO cells. Mutating either of the two LL motifs had a similar effect. We further assessed ASIC2a localization in organotypic hippocampal slice neurons. The LL motif mutants exhibited increased dendritic trafficking and elevated targeting to dendritic spines. Consistent with an efficient trafficking, the LL motif mutants increased acid-activated current density. In addition, mutating the second LL motif increased pH sensitivity of the channel. These data identify the LL motifs as a negative regulator of ASIC2a trafficking and function, and suggest novel regulatory mechanisms in acid signaling.

Cultured neuronal cell lines can express properties of mature neurons if properly differentiated. Although the precise mechanisms underlying neuronal differentiation are not fully understood, the expression and activation of ion channels, particularly those of Ca2+-permeable channels, have been suggested to play a role. In this study, we explored the presence and characterized the properties of acid-sensing ion channels (ASICs) in NS20Y cells, a neuronal cell line previously used for the study of neuronal differentiation. In addition, the potential role of ASICs in cell differentiation was explored. Reverse Transcription Polymerase Chain Reaction and Western blot revealed the presence of ASIC1 subunits in these cells. Fast drops of extracellular pH activated transient inward currents which were blocked, in a dose dependent manner, by amiloride, a non-selective ASIC blocker, and by Psalmotoxin-1 (PcTX1), a specific inhibitor for homomeric ASIC1a and heteromeric ASIC1a/2b channels. Incubation of cells with PcTX1 significantly reduced the differentiation of NS20Y cells induced by cpt-cAMP, as evidenced by decreased neurite length, dendritic complexity, decreased expression of functional voltage gated Na+ channels. Consistent with ASIC1a inhibition, ASIC1a knockdown with small interference RNA significantly attenuates cpt-cAMP-induced increase of neurite outgrowth. In summary, we described the presence of functional ASICs in NS20Y cells and demonstrate that ASIC1a plays a role in the differentiation of these cells.