ATP-sensitive K؉ (K ATP ) channels may be regulated by protons in addition to ATP, phospholipids, and other nucleotides. Such regulation allows a control of cellular excitability in conditions when pH is low but ATP concentration is normal. However, whether the K ATP changes its activity with pH alterations remains uncertain. In this study we showed that the reconstituted K ATP was strongly activated during hypercapnia and intracellular acidosis using whole-cell recordings. Further characterizations in excised patches indicated that channel activity increased with a moderate drop in intracellular pH and decreased with strong acidification. The channel activation was produced by a direct action of protons on the Kir6 subunit and relied on a histidine residue that is conserved in all K ATP . The inhibition appeared to be a result of channel rundown and was not seen in whole-cell recordings. The biphasic response may explain the contradictory pH sensitivity observed in cell-endogenous K ATP in excised patches. Site-specific mutations of two residues showed that pH and ATP sensitivities were independent of each other. Thus, these results demonstrate that the proton is a potent activator of the K ATP . The pH-dependent activation may enable the K ATP to control vascular tones, insulin secretion, and neuronal excitability in several pathophysiologic conditions. Hypercapnia and acidosis affect vascular tone, skeletal muscle contractility, insulin secretion, epithelial transport, and neuronal excitability, which may be mediated by K ATP 1 (1-5). However, previous studies on the pH sensitivity of these K ϩ channels were controversial and even contradictory. In the absence of ATP, acidic pH was shown to stimulate cell-endogenous K ATP (6, 7), inhibit it (8, 9), and have little or no effect (10, 11). This inconsistency is further complicated by the indirect effect of ATP or Mg 2ϩ and tissue-specific K ATP species (8 -12). Consequently, it is unclear whether K ATP is modulated during hypercapnia and acidosis and what molecular mechanisms are underlying the modulations. The cloned K ATP channels are ideal for addressing these questions because they allow for fine dissection of the modulatory mechanisms and elaborate manipulation of PCO 2 and pH in an expression system (13,14). Therefore, we studied the modulation of the cloned K ATP (Kir6 with SUR, Ref. 15) by CO 2 and acidic pH. To locate the pH sensors, we also studied Kir6.2 with a truncation of 36 amino acids at the C terminus (Kir6.2⌬C36) because it expresses functional channel without the SUR subunit and retains fair ATP sensitivity (16). MATERIALS AND METHODSOocytes from Xenopus laevis were used in the present studies. Frogs were anesthetized by bathing them in 0.3% 3-aminobenzoic acid ethyl ester. A few lobes of ovaries were removed after a small abdominal incision (ϳ5 mm). Then, the surgical incision was closed and the frogs were allowed to recover from the anesthesia. Xenopus oocytes were treated with 2 mg/ml collagenase (Type I, Sigma) in an OR2 solution consistin...

Investigating the mechanistic influence of the tumor microenvironment on cancer cell migration and membrane blebbing is crucial in the understanding and eventual arrest of cancer metastasis. In this study, we investigate the effect of suspended and aligned nanofibers on the glioma cytoskeleton, cell shape, migration and plasma membrane blebbing dynamics using a non-electrospinning fiber-manufacturing platform. Cells attached in repeatable shapes of spindle on single fibers, rectangular on two parallel fibers and polygonal on intersecting fibers. Structural stiffness (N m(-1)) of aligned and suspended nanofibers (average diameter: 400 nm, length: 4, 6, and 10 mm) was found to significantly alter the migration speed with higher migration on lower stiffness fibers. For cells attached to fibers and exhibiting blebbing, an increase in cellular spread area resulted in both reduced bleb count and bleb size with an overall increase in cell migration speed. Blebs no longer appeared past a critical cellular spread area of approximately 1400 μm(2). Our results highlighting the influence of the mechanistic environment on the invasion dynamics of glioma cells add to the understanding of how biophysical components influence glioma cell migration and blebbing dynamics.

Cell migration is studied with the traditional focus on protrusion-driven cell body displacement, while less is known on morphodynamics of individual protrusions themselves, especially in fibrous environments mimicking extracellular matrix. Here, using suspended fibers, we report integrative and multiscale abilities to study protrusive behavior independent of cell body migration. By manipulating the diameter of fibers in orthogonal directions, we constrain cell migration along large diameter (2 μm) base fibers, while solely allowing cells to sense, initiate, and mature protrusions on orthogonally deposited high-curvature/low diameter (∼100, 200, and 600 nm) protrusive fibers and low-curvature (∼300 and 600 nm width) protrusive flat ribbons. In doing so, we report a set of morphodynamic metrics that precisely quantitate protrusion dynamics. Protrusion growth and maturation occur by rapid broadening at the base to achieve long lengths, a behavior dramatically influenced by curvature. While flat ribbons universally induce the formation of broad and long protrusions, we quantitatively protrutype protrusive behavior of two highly invasive cancer cell lines and find breast adenocarcinoma (MDA-MB-231) to exhibit sensitivity to fiber curvature higher than that of brain glioblastoma DBTRG-05MG. Furthermore, while actin and microtubules localize within protrusions of all sizes, we quantify protrusion size-driven localization of vimentin and, contrary to current understanding, report that vimentin is not required to form protrusions. Using multiple protrusive fibers, we quantify high coordination between hierarchical branches of individual protrusions and describe how the spatial configuration of multiple protrusions regulates cell migratory state. Finally, we describe protrusion-driven shedding and collection of cytoplasmic debris.

The novel coronavirus disease (COVID-19) has been declared by the World Health Organisation as an international public health emergency. Owing to its high infectivity, countries all over the world implemented nationwide lockdowns with the hope of flattening the epidemic curve. Around the world, this has led to the closure of schools in over 150 countries affecting the education of nearly 1 billion children (Sahu, 2020). India faced total lockdown from 24th March 2020 to May 2020 and even though a phased reopening of public services has since then been attempted, most educational institutions including schools and colleges remain closed without a clear view regarding their reopening. This has created an unprecedented crisis in the education sector for students as well as educators regarding continuation of educational services, conducting assessments and catering to the needs of special education and vocational rehabilitation. This paper discusses the various psychosocial issues that have emerged leading to academic stress amongst children and adolescent students and its potential to lead to short and long-term psychological morbidity.