Transient mitochondrial permeability transition pore opening mediates preconditioning-induced protection

Background - Transient (low-conductance) opening of the mitochondrial permeability transition pore (mPTP) may limit mitochondrial calcium load and mediate mitochondrial reactive oxygen species (ROS) signaling. We hypothesize that transient mPTP opening and ROS mediate the protection associated with myocardial preconditioning and mitochondrial uncoupling.Methods and Results - Isolated perfused rat hearts were subjected to 35 minutes of ischemia/ 120 minutes of reperfusion, and the infarct-risk-volume ratio was determined by tetrazolium staining. Inhibiting mPTP opening during the preconditioning phase with cyclosporine-A (CsA, 0.2 mumol/L) or sanglifehrin-A (SfA, 1.0 mumol/L) abolished the protection associated with ischemic preconditioning (IPC) ( 20.2 +/- 3.6% versus 45.9 +/- 2.5% with CsA, 49.0 +/- 7.1% with SfA; P < 0.001); and pharmacological preconditioning with diazoxide (Dzx, 30 mu mol/L) (22.1 +/- 2.7% versus 46.3 +/- 3.0% with CsA, 48.4 +/- 5.5% with SfA; P < 0.001), CCPA ( the adenosine A1-receptor agonist, 200 nmol/L) (24.9 +/- 4.5% versus 54.4 +/- 6.6% with CsA, 42.6 +/- 9.0% with SfA; P < 0.001), or 2,4-dinitrophenol (DNP, the mitochondrial uncoupler, 50 mu mol/L) (15.7 +/- 2.7% versus 40.8 +/- 5.5% with CsA, 34.3 +/- 3.1% with SfA; P < 0.001), suggesting that mPTP opening during the preconditioning phase is required to mediate protection in these settings. Inhibiting ROS during the preconditioning protocols with N-mercaptopropionylglycine (MPG, 1 mmol/L) also abolished the protection associated with IPC (20.2 +/- 3.6% versus 47.1 +/- 3.8% with MPG; P < 0.001), diazoxide (22.1 +/- 2.7% versus 56.3 +/- 3.8% with MPG; P < 0.001), and DNP (15.7 +/- 2.7% versus 50.7 +/- 6.6% with MPG; P < 0.001) but not CCPA (24.9 +/- 4.5% versus 26.5 +/- 8.4% with MPG; P = NS). Further experiments in adult rat myocytes demonstrated that diazoxide induced CsA-sensitive, low-conductance transient mPTP opening (represented by a 28 +/- 3% reduction in mitochondrial calcein fluorescence compared with control; P < 0.01).Conclusions - We report that the protection associated with IPC, diazoxide, and mitochondrial uncoupling requires transient mPTP opening and ROS

CLARIN: Common language resources and technology infrastructure

This paper gives an overview of the CLARIN project [1], which aims to create a research infrastructure that makes language resources and technology (LRT) available and readily usable to scholars of all disciplines, in particular the humanities and social sciences (HSS)

School District Leadership and Racial Justice: Examining the Use of a Culturally Responsive-Sustaining Framework to Advance Equity Between Black and White Student Populations Within New York State Public Education

The current climate in the United States reflects a heightened focus on examining and challenging systemic inequities that have long impacted Black individuals. Culturally responsive education efforts have increased as schools experience increasingly diverse populations. The purpose of this study was to gain an understanding of how district leaders are implementing the New York State Education Department (NYSED) Culturally Responsive-Sustaining (CR-S) Education Framework in advancing equity between their Black and White student populations. Challenges and barriers experienced in the implementation of this framework, and approaches and strategies needed to overcome these were also explored. Interviews with six school district leaders in New York State provided the sources for the data in this qualitative descriptive study. The results of the interviews reflect seven categories regarding the implementation of this framework. These categories include providing a starting point, professional development, the role of stakeholders, disproportionality, familiarity with the CR-S Framework, uncomfortable change, and diversity of action. Thirteen themes within these categories emerged from the interviews. Findings indicate a range of experiences with the implementation of the CR-S Education Framework, with school district leaders in the early phases of utilizing the framework to directly mitigate inequities between their Black and White student populations. The findings provide the basis for recommendations for policy development, improved practices, and further research needed in order to continue to advance educational equity throughout the public education system

Ethanol Sensitivity and Tolerance of Rat Neuronal BK Channels: A Dissertation

BK channels are well studied targets of acute ethanol action. They play a prominent role in neuronal excitability and have been shown to play a significant role in behavioral ethanol tolerance in invertebrates. The focus of my work centers on the effects of alcohol on the BK channel and comprises studies that examine how subcellular location affects acute ethanol sensitivity and how duration of acute alcohol exposure impacts the development of rapid tolerance. My results also provide potential mechanisms which underlie acute sensitivity and rapid tolerance. I first explore BK channel sensitivity to ethanol in the three compartments (dendrite, cell body, and nerve terminal) of magnocellular neurons in the rat hypothalamic-neurohypophysial (HNS) system. The HNS system provides a particularly powerful preparation in which to study the distribution and regional properties of ion channel proteins because the cell bodies are physically separated from the nerve terminals. Using electrophysiological and immunohistochemical techniques I characterize the BK channel in each of the three primary compartments and find that dendritic BK channels, similar to somatic channels, but in contrast to nerve terminal channels, are insensitive to alcohol. Furthermore, the gating kinetics, calcium sensitivity, and iberiotoxin sensitivity of channels in the dendrite are similar to somatic channels but sharply contrast terminal channels. The biophysical and pharmacological properties of somatodendritic vs. nerve terminal channels are consistent with the characteristics of exogenously expressed αβ1 vs. αβ4 channels, respectively. Therefore, one possible explanation for my findings is a selective distribution of β1 subunits to the somatodendritic compartment and β4 subunits to the terminal compartment. This hypothesis is supported immunohistochemically by the appearance of distinct punctate β1 or β4 channel clusters in the membrane of somatodendritic or nerve terminal compartments, respectively. In conclusion, I found that alcohol sensitivity of BK channels within the HNS system is dependent on subcellular location and postulate that β-subunits modulate ethanol sensitivity of HNS BK channels. In the second and primary focus of my thesis I explore tolerance development in the striatum, a brain region heavily implicated in addiction. Numerous studies have demonstrated that duration of drug exposure influences tolerance development and drug dependence. To further elucidate the mechanisms underlying behavioral tolerance I examined if BK channel tolerance was dependent on duration of alcohol exposure using patch clamp techniques in cultured striatal neurons from P8 rats. I found that persistence of rapid tolerance is indeed a function of exposure time and find it lasts surprisingly long. For example, after a 6 hr exposure to 20 mM ethanol, acute sensitivity was still suppressed at 24 hrs withdrawal. However, after a 1 or 3 hr exposure period, sensitivity had returned after only 4 hrs. I also found that during withdrawal from a 6 hr but not a 3 hr exposure the biophysical properties of BK channels change and that this change is correlated with an increase in mRNA levels of the alcohol insensitive STREX splice variant. Furthermore, BK channel properties during withdrawal from a 6 hr exposure to alcohol closely parallel the properties of STREX channels exogenously expressed in HEK293 cells. In conclusion I have established that BK channels develop rapid tolerance in striatal neurons, that rapid tolerance is dependent upon exposure protocol, and is surprisingly persistent. These findings present another mechanism underlying BK channel tolerance and possibly behavioral tolerance. Since these phenomena are dependent on duration of drug exposure my results may find relevance in explaining how drinking patterns impact the development of alcohol dependence in humans

Localising discrete points in 3D space using stereo pairs of digital slot-scanning X-rays

Includes bibliographical references (p. 104-111)