Carbonic Anhydrase-8 Regulates Inflammatory Pain by Inhibiting the ITPR1-Cytosolic Free Calcium Pathway

Calcium dysregulation is causally linked with various forms of neuropathology including seizure disorders, multiple sclerosis, Huntington’s disease, Alzheimer’s, spinal cerebellar ataxia (SCA) and chronic pain. Carbonic anhydrase-8 (Car8) is an allosteric inhibitor of inositol trisphosphate receptor-1 (ITPR1), which regulates intracellular calcium release fundamental to critical cellular functions including neuronal excitability, neurite outgrowth, neurotransmitter release, mitochondrial energy production and cell fate. In this report we test the hypothesis that Car8 regulation of ITPR1 and cytoplasmic free calcium release is critical to nociception and pain behaviors. We show Car8 null mutant mice (MT) exhibit mechanical allodynia and thermal hyperalgesia. Dorsal root ganglia (DRG) from MT also demonstrate increased steady-state ITPR1 phosphorylation (pITPR1) and cytoplasmic free calcium release. Overexpression of Car8 wildtype protein in MT nociceptors complements Car8 deficiency, down regulates pITPR1 and abolishes thermal and mechanical hypersensitivity. We also show that Car8 nociceptor overexpression alleviates chronic inflammatory pain. Finally, inflammation results in downregulation of DRG Car8 that is associated with increased pITPR1 expression relative to ITPR1, suggesting a possible mechanism of acute hypersensitivity. Our findings indicate Car8 regulates the ITPR1-cytosolic free calcium pathway that is critical to nociception, inflammatory pain and possibly other neuropathological states. Car8 and ITPR1 represent new therapeutic targets for chronic pain.     

Evaluating the Return in Ecosystem Services from Investment in Public Land Acquisitions

We evaluate the return on investment (ROI) from public land conservation in the state of Minnesota, USA. We use a spatially-explicit modeling tool, the Integrated Valuation of Ecosystem Services and Tradeoffs (InVEST), to estimate how changes in land use and land cover (LULC), including public land acquisitions for conservation, influence the joint provision and value of multiple ecosystem services. We calculate the ROI of a public conservation acquisition as the ratio of the present value of ecosystem services generated by the conservation to the cost of the conservation. For the land scenarios analyzed, carbon sequestration services generated the greatest benefits followed by water quality improvements and recreation opportunities. We found ROI values ranged from 0.21 to 5.28 depending on assumptions about future land use change, service values, and discount rate. Our study suggests conservation is a good investment as long as investments are targeted to areas with low land costs and high service values.     

When is now? Perception of simultaneity.

We address the following question: Is there a difference (D) between the amount of time for auditory and visual stimuli to be perceived? On each of 1000 trials, observers were presented with a light-sound pair, separated by a stimulus onset asynchrony (SOA) between -250 ms (sound first) and +250 ms. Observers indicated if the light-sound pair came on simultaneously by pressing one of two (yes or no) keys. The SOA most likely to yield affirmative responses was defined as the point of subjective simultaneity (PSS). PSS values were between -21 ms (i.e. sound 21 ms before light) and +150 ms. Evidence is presented that each PSS is observer specific. In a second experiment, each observer was tested using two observer-stimulus distances. The resultant PSS values are highly correlated (r = 0.954, p = 0.003), suggesting that each observer''s PSS is stable. PSS values were significantly affected by observer-stimulus distance, suggesting that observers do not take account of changes in distance on the resultant difference in arrival times of light and sound. The difference RTd in simple reaction time to single visual and auditory stimuli was also estimated; no evidence that RTd is observer specific or stable was found. The implications of these findings for the perception of multisensory stimuli are discussed.     

The kinetics of binding human prolactin, but not growth hormone, to the prolactin receptor vary over a physiologic pH range

A member of the family of hematopoietic cytokines, human prolactin (hPRL) serves a dual role both as an endocrine hormone and as an autocrine/paracrine cytokine or growth factor. During investigation of the solution structural properties of hPRL, we have noted a surprising pH dependence of its structural stability over a range from approximately pH 6.0 to pH 8.0. An analysis of backbone atom NMR chemical shift changes and backbone amide hydrogen-deuterium exchange rates due to titration of the solution pH over this same range, along with calculations of protein surface electrostatic potential, suggests the possible involvement of a localized cluster of three His residues (27, 30, and 180), which comprise a portion of the high-affinity receptor-binding epitope. Surface plasmon resonance analysis of the interaction between hPRL and the extracellular domain (ECD) of the hPRL receptor reveals a selective 500-fold change in the dissociation rate between pH 8.3 and pH 5.8. In comparison, the interaction of hGH with the same receptor ECD did not demonstrate any significant dependence on pH. We also present an initial investigation of the pH dependence of hPRL function in rat Nb2 cell proliferation assays and a STAT5 luciferase gene reporter assay in the T47D human breast cancer cell line, whose results are consistent with our biophysical studies. The potential implications of this variation in hPRL's structural stability and receptor-binding kinetics over this physiologic range of pH are discussed.     

Abnormal renal response to twenty-four-hour dehydration and fasting in Nagase analbuminemic rats.

We assessed renal function in fasting adult Nagase analbuminemic rats (NAR). Sodium output in male and female NAR was 68% and 46%, respectively, of the output of age- and sex-matched normal Sprague-Dawley (SD) rats. Potassium excretion was significantly greater in female NAR but there was no difference between male NAR and SD rats. The renal clearances of urea and creatinine were reduced in NAR with corresponding increases in plasma concentrations; however, the urea and creatinine concentrations were not different in plasma samples taken from normally fed and hydrated SD and NAR rats. Exchangeable body sodium and sodium space was significantly larger in normally fed and hydrated NAR than in SD but there were no differences in plasma sodium concentrations or plasma volumes. Although plasma concentrations of albumin in NAR were only about 0.07% of the concentration in SD rats, the renal clearance of albumin in NAR was threefold greater. Kidney weights in NAR were 10 to 16% less than in SD rats but liver weights were 22 to 42% greater. Clearly, renal function was markedly abnormal in Nagase rats during a 24-hour fast.     

Coxsackievirus and adenovirus receptor is essential for cardiomyocyte development

The coxsackievirus and adenovirus receptor (CAR) is a transmembrane protein that is known to be a site of viral attachment and entry, but its physiologic functions are undefined. CAR expression is maximal in neonates and wanes rapidly after birth in organs such as heart, muscle, and brain, suggesting that CAR plays a role in the development of these tissues. Here, we show that CAR deficiency resulted in an embryonic lethal condition associated with cardiac defects. Specifically, commencing approximately 10.5 days postconception (dpc), CAR-/- cardiomyocytes exhibited regional apoptosis evidenced by both histopathologic features of cell death and positive staining for the apoptotic marker cleaved caspase 3. CAR-/- fetuses invariably suffered from degeneration of the myocardial wall and thoracic hemorrhaging, leading to death by 11.5 dpc. These findings are consistent with the view that CAR provides positive survival signals to cardiomyocytes that are essential for normal heart development.