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Pain is both a symptom and a disease. It manifests in multiple forms and its treatment is complex. Physical, social, economic, and emotional consequences of pain can impair an individual's overall health, well-being, productivity, and relationships in myriad ways. The impact of pain at a population level is vast and, while estimates differ, the Centers for Disease Control and Prevention reported that 50 million U.S. adults are living in pain. In terms of pain's global impact, estimates suggest the problem affects approximately 1 in 5 adults across the world, with nearly 1 in 10 adults newly diagnosed with chronic pain each year. In recent years, the issues surrounding the complexity of pain management have contributed to increased demand for alternative strategies for treating pain. One such strategy is to expand use of topical pain medications—medications applied to intact skin. This nonoral route of administration for pain medication has the potential benefit, in theory, of local activity and fewer systemic side effects. Compounding is an age-old pharmaceutical practice of combining, mixing, or adjusting ingredients to create a tailored medication to meet the needs of a patient. The aim of compounding, historically, has been to provide patients with access to therapeutic alternatives that are safe and effective, especially for people with clinical needs that cannot otherwise be met by commercially available FDA-approved drugs. Compounded Topical Pain Creams explores issues regarding the safety and effectiveness of the ingredients in these pain creams. This report analyzes the available scientific data relating to the ingredients used in compounded topical pain creams and offers recommendations regarding the treatment of patients.

Medical education has undergone a wave of creative innovation over the last decade, with new curricular structures, pedagogy, content, and team-based approaches. Augmenting these changes, integration of clinical and scientific principles increasingly occurs across all years of training. Given success in innovation and integration, as well as recent interest and national pilots in time-variable (competency-based) education, we propose the next important step in medical education evolution is individualization.

Christopher D Stamy,1 Christine C Schwartz,1 Danielle A Phillips,2 Aparna S Ajjarapu,3 Kristi J Ferguson,4,5 Debra A Schwinn6–8 1University of Iowa Carver College of Medicine, Iowa City, 2Des Moines University Osteopathic Medical Center, Des Moines, 3University of Iowa, 4Office of Consultation& Research in Medical Education, University of Iowa Carver College of Medicine, 5Department of Internal Medicine, 6Department of Anesthesia, 7Department of Biochemistry, 8Department of Pharmacology, University of Iowa Health Care, Iowa City, IA, USA Background: Medical education is undergoing robust curricular reform with several innovative models emerging. In this study, we examined current trends in 3-year Doctor of Medicine (MD) education and place these programs in context. Methods: A survey was conducted among Deans of U.S. allopathic medical schools using structured phone interview regarding current availability of a 3-year MD pathway, and/or other variations in curricular innovation, within their institution. Those with 3-year programs answered additional questions. Results: Data from 107 institutions were obtained (75% survey response rate). The most common variation in length of medical education today is the accelerated 3-year pathway. Since 2010, 9 medical schools have introduced parallel 3-year MD programs and another 4 are actively developing such programs. However, the total number of students in 3-year MD tracks remains small (n=199 students, or 0.2% total medical students). Family medicine and general internal medicine are the most common residency programs selected. Benefits of 3-year MD programs generally include reduction in student debt, stability of guaranteed residency positions, and potential for increasing physician numbers in rural/underserved areas. Drawbacks include concern about fatigue/burnout, difficulty in providing guaranteed residency positions, and additional expense in teaching 2 parallel curricula. Four vignettes of alternative innovative and relevant curricular initiatives are also presented in order to place 3-year MD programs in a broader context of medical education reform in the U.S. Conclusion: Three-year MD pathways are the most common accelerated alternative available at a small number of medical schools for highly selected students. Long-term evaluation of these programs will be essential to determine if these programs are meeting their goals (e.g., increasing the number of physicians in rural/underserved areas). Benefits and shortcomings of such programs should be carefully examined when considering this approach, or others described, as part of MD curricular options designed to individualize medical education. Keywords: pedagogy, curriculum, 3-year MD, medical students, medical school, reform

The role of naturally occurring human α1a-Adrenergic Receptor (α1aAR) genetic variants associated with cardiovascular disorders is poorly understood. Here, we present the novel findings that expression of human α1aAR-247R (247R) genetic variant in cardiomyoblasts leads to transition of cardiomyoblasts into a fibroblast-like phenotype, evidenced by morphology and distinct de novo expression of characteristic genes. These fibroblast-like cells exhibit constitutive, high proliferative capacity and agonist-induced hypertrophy compared with cells prior to transition. We demonstrate that constitutive, synergistic activation of EGFR, Src and ERK kinases is the potential molecular mechanism of this transition. We also demonstrate that 247R triggers two distinct EGFR transactivation-dependent signaling pathways: 1) constitutive Gq-independent β-arrestin-1/Src/MMP/EGFR/ERK-dependent hyperproliferation and 2) agonist-induced Gq- and EGFR/STAT-dependent hypertrophy. Interestingly, in cardiomyoblasts agonist-independent hyperproliferation is MMP-dependent, but in fibroblast-like cells it is MMP-independent, suggesting that expression of α1aAR genetic variant in cardiomyocytes may trigger extracellular matrix remodeling. Thus, these novel findings demonstrate that EGFR transactivation by α1aAR-247R leads to hyperproliferation, hypertrophy and alterations in cardiomyoblasts, suggesting that these unique genetically-mediated alterations in signaling pathways and cellular function may lead to myocardial fibrosis. Such extracellular matrix remodeling may contribute to the genesis of arrhythmias in certain types of heart failure.

The purpose of this report is to describe student satisfaction with a near-peer interprofessional education (IPE) session for physical therapy and medical students. Ten senior physical therapy students worked in peer-groups to develop a musculoskeletal anatomy demonstration for first-semester medical students. Together with their classmates, they demonstrated observation, palpation, and musculoskeletal assessment of the shoulder and scapular-thoracic articulation to medical student dissection groups in the Gross Anatomy laboratory. The medical students were encouraged to consider the synergistic function of shoulder structures and the potential impact of a selected pathology: rotator cuff injury. The session provided the medical students with an opportunity to integrate their new anatomical knowledge into a framework for clinical musculoskeletal evaluation. The experience offered senior physical therapy students an opportunity to work in teams with their peers, internalize and adapt to constructive feedback, and seek common ground with members of another profession. Both student groups reported a high degree of satisfaction with the sessions and expressed a desire for further interaction. These positive perceptions by student stakeholders have prompted us to consider additional IPE exchanges for the anatomy course in the upcoming school year. Given the positive outcome of this descriptive study, we now plan to systematically test whether near-peer IPE interactions can enhance the degree that students learn key anatomical concepts.

Genomic variation is an important factor in why supposedly “similar” patients react differently to drugs, have different disease course(s), and varying clinical outcomes. This review provides an update on concepts in modern genomic medicine with an emphasis on clinically relevant study approaches, disease/drug pathway analysis, and recent pharmacogenomic findings. The application of genomic medicine and its importance for rapid diagnosis of disease-causing agents, as well as its clinical application in human disease diagnosis/treatment and in cardiovascular disease are discussed. In addition to direct clinical applications, modern genomic approaches also play an important role in elucidating new mechanisms of disease. Finally, the role of the National Institutes of Health national pharmacogenomics research network in codifying “bench to bedside” translation of genetic results that impact drug therapy will also be discussed.

α 1- adrenergic receptors (α 1a , α 1b , α 1d ), members of G protein-coupled receptor (GPCR) family, regulate stress-facilitated changes in the cardiovascular (CV) system and human blood pressure (BP) via smooth muscle cell (SMC) proliferation and vasoconstriction. The α 1a AR is the predominant subtype in human heart and resistance vessels. The α 1a AR G247R (247R) genetic variant was discovered in a patient with severe hypertension and in a cohort of patients enriched for CV disorders. We reported a unique coupling of this genetic variant to βarrestin/MMP/EGFR transactivation pathway in rat fibroblasts, cardiomyoblasts and vascular SMCs leading to hyperproliferation and hypertrophy. Here we present that differential interaction of 247R or α 1a AR (WT) with scaffolding/regulatory proteins mediate unique signaling of 247R. Physiological levels of 247R but not WT upregulate endogenous levels of scaffolding protein spinophilin (SPL) in rat SMCs and cardiomyoblasts. SPL knockdown or overexpression of RGS2 (negative R egulators of G -protein S ignaling) inhibits 247R-mediated hyperproliferation. Overexpression of SPL with RGS2 restores hyperproliferation suggesting that SPL binds RGS2 and prevents 247R-RGS2 interaction. To determine if 247R hyperproliferation is in effect in human cells, we generated human iPSCs from primary fibroblasts and differentiated them to SMCs with characteristic morphology and expression of SMC markers (RT-PCR at various passages). Consistent with our findings in rat cells, expression of 247R but not WT in hiPSC-derived SMCs triggered ~70% increased proliferation. Agonist treatment resulted in increased hypertrophy in 247R cells compared with WT. Co-immunoprecipitation from HEK293 cells expressing HA-α 1 ARs and full length Myc-SPL or Flag-RGS2 revealed that SPL exhibits ~2-fold stronger interaction with WT than with 247R. Consequently, SPL recruits RGS2 and inhibits receptor signaling. In contrast, weaker SPL-247R interaction results in diminished RGS2 inhibitory effect allowing hyperproliferation of 247R cells. Thus, we identify novel SPL/RGS2 signalosome responsible for α 1a AR signaling in human cells and identify SPL as a potential novel target for treatment of α 1 AR-mediated human CV disorders.

We previously identified a naturally occurring human SNP, G247R, in the third intracellular loop of the α 1a -adrenergic receptor (α 1a -247R) and demonstrated that constitutive expression of α 1a -247R results in twofold increased cell proliferation compared with WT. In the present study we elucidate molecular mechanisms and signal transduction pathways responsible for increased cell proliferation unique to α 1a -247R, but not α 1a -WT, α 1b , or α 1d AR subtypes. We show that elevated levels of matrix metalloproteinase-7 (MMP7) and a disintegrin and metalloproteinase-12 (ADAM12) in α 1a -247R–expressing cells are responsible for EGF receptor (EGFR) transactivation, downstream ERK activation, and increased cell proliferation; this pathway is confirmed using MMP, EGFR, and ERK inhibitors. We demonstrate that EGFR transactivation and downstream ERK activation depends on increased shedding of heparin-binding EGF. Finally, we demonstrate that knockdown of MMP7 or β-arrestin1 by shRNAs results in attenuation of proliferation of cells expressing α 1a -247R. Importantly, accelerated cell proliferation triggered by the α 1a -247R is serum- and agonist-independent, providing unique evidence for constitutive active coupling to the β-arrestin1/MMP/EGFR transactivation pathway by any G protein-coupled receptor. These findings raise the possibility of a previously unexplored mechanism for sympathetically mediated human hypertension triggered by a naturally occurring human genetic variant.

Objective Hemorrhage continues to be a major problem after cardiac surgery despite the routine use of antifibrinolytic drugs, with striking inter-patient variability poorly explained by already known risk factors. The authors tested the hypothesis that genetic polymorphisms of inflammatory mediators and cellular adhesion molecules are associated with bleeding after cardiac surgery. Design Prospective, observational study. Setting Single, tertiary referral university heart center. Participants Adult patients undergoing aortocoronary surgery with cardiopulmonary bypass. Interventions Patients (n = 759) had 10 mL of blood drawn preoperatively and genomic DNA isolated then genotyped for 17 polymorphisms in 7 candidate genes: tumor necrosis factor, interleukins 1β and 6, interleukin 1 receptor antagonist, intercellular adhesion molecule-1 (ICAM-1), P-selectin and endothelial leucocyte adhesion molecule-1 (E-selectin). Multivariate analyses were used to relate clinical and genetic factors to bleeding and transfusion. Measurements and Main Results The 98G/T polymorphism of the E-selectin gene was independently associated with bleeding after cardiac surgery (p = 0.002), after adjusting for significant clinical predictors (patient size and baseline hemoglobin concentration). There was a gene dose effect according to the number of minor alleles in the genotype; carriers of the minor allele bled 17% (GT) and 54% (TT) more than wild type (GG) genotypes, respectively (p = 0.01). Carriers of the minor allele also had longer activated partial thromboplastin times (p = 0.0023) and increased fresh frozen plasma transfusion (p = 0.03) compared with wild type. Conclusions The authors found a dose-related association between the 98T E-selectin polymorphism and bleeding after cardiac surgery, independent of and additive to standard clinical risk factors.

We have reported that the alpha(1A)-adrenergic receptor (alpha(1A)AR) in rat-1 fibroblasts is a lipid raft protein. Here we examined whether disrupting lipid rafts by methyl-beta-cyclodextrin (MCD) sequestration of cholesterol affects alpha(1A)AR signaling. Unexpectedly, MCD increased alpha(1A)AR-dependent basal inositol phosphate formation and p38 mitogen-activated protein kinase activation in a cholesterol-dependent manner. It also initiated internalization of surface alpha(1A)AR, which was partially blocked by receptor inhibition. Binding assays revealed MCD-mediated increases in receptor agonist affinity as well as reciprocal decreases in inverse agonist affinity, a behavior that is usually interpreted as a shift toward the active receptor conformation. In untreated cells a fraction of the receptor was found to be present in preassociated receptor/G protein complexes, which rapidly dissociate upon receptor stimulation. Consistent with MCD-induced signaling, raft disruption resulted in an increase in receptor/G protein complexes. These results strongly suggest that lipid rafts constrain basal alpha(1A)AR activity; however, preassembled receptor/G protein complexes could still provide a mechanism for accelerating alpha(1A)AR signaling following stimulation.

Genetic variation plays an important role in chronic disease and in perioperative patient management. It was our intent in this short review to cite practical examples of relevant overlap between pharmacoge-nomics and clinical anesthesiology. The influence of genetics on patient safety is seen throughout the perioperative period, when evaluating patients with co-existing diseases in the pre-anesthesia clinic, when considering metabolism and efficacy of drugs intra-operatively, and at the time of postoperative pain management and clinical outcome assessment. The situations presented in this editorial provide compel-ling reasons for clinicians to embrace the partnership between genetics and anesthesiology.

It is unclear whether the regulatory distinction between non-identifiable and identifiable information--information used to determine informed consent practices for the use of clinically derived samples for genetic research--is meaningful to patients. The objective of this study was to examine patients' attitudes and preferences regarding use of anonymous and identifiable clinical samples for genetic research. Telephone interviews were conducted with 1,193 patients recruited from general medicine, thoracic surgery, or medical oncology clinics at five United States academic medical centers. Wanting to know about research being done was important to 72% of patients when samples would be anonymous and to 81% of patients when samples would be identifiable. Only 17% wanted to know about the identifiable scenario but not the anonymous scenario (i.e., following the regulatory distinction). Curiosity-based reasons were the most common (37%) among patients who wanted to know about anonymous samples. Of patients wanting to know about either scenario, approximately 57% would require researchers to seek permission, whereas 43% would be satisfied with notification only. Patients were more likely to support permission (versus notification) in the anonymous scenario if they had more education, were Black, less religious, in better health, more private, and less trusting of researchers. The sample, although not representative of the general population, does represent patients at academic medical centers whose clinical samples may be used for genetic research. Few patients expressed preferences consistent with the regulatory distinction between non-identifiable and identifiable information. Data from this study should cause policy-makers to question whether this distinction is useful in relation to research with previously collected clinically derived samples.

Genomic medicine has provided new mechanistic understanding for many complex diseases over the last 5–10 years. More recently genomic approaches have been applied to the perioperative paradigm, facilitating identification of patients at high risk for adverse events, as well as those who will respond better/worse to specific pharmacologic therapies. The consistent biological theme emerging is that while inflammation is important in healing from surgical trauma, patients who are too robustly proinflammatory appear to be at higher risk for adverse perioperative events. Precise predictors of each adverse event are being elucidated so that corrective therapeutics can be instituted to improve outcomes in high-risk patients. While the field of perioperative genomics could be considered in its infancy, such approaches are the wave of the future.

Benign prostatic hyperplasia (BPH) is a common cause of urinary outflow obstruction in aging men leading to lower urinary tract symptoms (LUTS). α1-Adrenoceptors (α1ARs) antagonists (blockers) have become a mainstay of LUTS treatment because they relax prostate smooth muscle and decrease urethral resistance, as well as relieving bladder LUTS symptoms. A review of key recent clinical trials suggests new insights into the role of specific α1AR subtypes in the treatment of LUTS.

The alpha(1a)-adrenergic receptor (alpha(1a)AR) occupies intracellular and plasma membranes in both native and heterologous expression systems. Based on multiple independent lines of evidence, we demonstrate the alpha(1a)AR at the cell surface occupies membrane rafts but exits from rafts following stimulation. In non-detergent raft preparations, basal alpha(1a)AR is present in low density membrane rafts and colocalizes with its G protein effectors on density gradients. Raft disruption by cholesterol depletion with methyl-beta-cyclodextrin eliminates these light rafts. To confirm the presence of the alpha(1a)AR in plasma membrane rafts, fluorescence resonance energy transfer measurements were used to demonstrate colocalization of surface receptor and the raft marker, cholera toxin B. This colocalization was largely lost following alpha(1a)AR stimulation with phenylephrine. Similarly, receptor stimulation causes exit of the alpha(1a)AR from light rafts within 3-10 min in contrast to the G proteins, which largely remain in light rafts. Importantly, this delayed exit of the alpha(1a)AR suggests acute receptor signaling and desensitization occur entirely within rafts. Interestingly, both confocal analysis and measurement of surface alpha(1a)AR levels indicate modest receptor internalization during the 10 min following stimulation, suggesting most of the receptor has entered non-raft plasma membrane. Nevertheless, activation does increase the rate of receptor internalization as does disruption of rafts with methyl-beta-cyclodextrin, suggesting raft exit enables internalization. Confocal analysis of surface-labeled hemagglutinin-alpha(1a)AR reveals that basal and stimulated receptor occupies clathrin pits in fixed cells consistent with previous indirect evidence. The evidence presented here strongly suggests the alpha(1a)AR is a lipid raft protein under basal conditions and implies agonist-mediated signaling occurs from rafts.

A growing body of evidence implicates alpha1-adrenergic receptors (alpha1ARs) as potent regulators of growth pathways. The three alpha1AR subtypes (alpha1aAR, alpha1bAR, alpha1dAR) display highly restricted tissue expression that undergoes subtype switching with many pathological stimuli, the mechanistic basis of which remains unknown. To gain insight into transcriptional pathways governing cell-specific regulation of the human alpha1dAR subtype, we cloned and characterized the alpha1dAR promoter region in two human cellular models that display disparate levels of endogenous alpha1dAR expression (SK-N-MC and DU145). Results reveal that alpha1dAR basal expression is regulated by Sp1-dependent binding of two promoter-proximal GC boxes, the mutation of which attenuates alpha1dAR promoter activity 10-fold. Mechanistically, chromatin immunoprecipitation data demonstrate that Sp1 binding correlates with expression of the endogenous gene in vivo, correlating highly with alpha1dAR promoter methylation-dependent silencing of both episomally expressed reporter constructs and the endogenous gene. Further, analysis of methylation status of proximal GC boxes using sodium bisulfite sequencing reveals differential methylation of proximal GC boxes in the two cell lines examined. Together, the data support a mechanism of methylation-dependent disruption of Sp1 binding in a cell-specific manner resulting in repression of basal alpha1dAR expression.

Objectives : Human α 1 adrenergic receptors (AR), members of G protein-coupled receptor superfamily (GPCR) regulate blood pressure via smooth muscle cell (SMC) proliferation and vasoconstriction. We showed that α 1a AR-G247R (247R) genetic variant, identified in a hypertensive patient, constitutively couples to β-arrestin1/MMP/EGFR transactivation pathway leading to hyperproliferation in fibroblasts, cardiomyoblasts and SMCs. A scaffolding protein spinophilin (SPL) serves as a docking site for many regulatory proteins including RGS2 (negative R egulators of G -protein S ignaling). SPL also binds 3 rd intracellular loop of some GPCRs. Hypothesis: differential interaction of 247R or α 1a AR-WT (WT) with SPL/RGS2/β-arrestin signalosome mediates unique signaling of 247R and hyperproliferation in cardiovascular cells. Methods: Receptor-protein interactions were determined by co-immunoprecipitation from HEK293 cells expressing HA-α 1 ARs and full length Myc-SPL, its fragments or Flag-RGS2. Protein levels were analyzed by Western. SPL knockdown or RGS2 overexpression was performed using SPL-specific or scrambled siRNA or Flag-RGS2. Results: Our results reveal that in SMCs or cardiomyoblasts 247R but not WT upregulates endogenous SPL. SPL exhibits stronger (~2-fold) interaction with WT compared to 247R or α 1b , recruits RGS2 and inhibits receptor signaling. In contrast, weak SPL-247R interaction diminishes RGS2 inhibitory effect permitting hyperproliferation of 247R cells. SPL knockdown inhibits 247R-induced proliferation by allowing RGS2 directly bind to 247R as observed with RGS2 overexpression. Overexpression of SPL with RGS2 restores hyperproliferation suggesting that in 247R cells SPL binds RGS2 preventing RGS2-247R interaction. Conclusions: We present SPL/RGS2/β-arrestin as a novel signalosome responsible for α 1a AR-247R genetic variant triggered hyperproliferation in different cardiovascular cells. We reveal that SPL regulates α 1a AR signaling by differentially binding WT or 247R receptors and recruiting RGS2 protein to the receptor. These novel findings unravel critical roles of SPL and RGS2 in α 1 AR signaling, as well as identify SPL as a potential novel target for treatment of α 1 AR-mediated cardiovascular disorders.

Reductions in federal support and clinical revenue jeopardize biomedical research and, in turn, clinical medicine.

Physician scientists perform biomedical research ranging from fundamental molecular studies to clinical trials. As such, physician scientists have a pivotal role in the biomedical research enterprise. The foundation for physician scientist leadership in the subspecialty of anesthesiology is highly dependent on the success of young investigators as they develop independent careers involving basic and clinical investigation. Hence, a leading priority of anesthesiology must be to nurture the academic careers of physician scientists within the specialty. Over the past decade, during a period of budgetary expansion by the National Institutes of Health (NIH), the number of NIH grant applications from physicians, in contrast to the number of grant proposals from Ph.D. scientists, has been slow to increase. Likewise, growth in NIH awards to physicians working in anesthesiology departments has also been slow.1 Junior clinical anesthesiology faculty across the country find their ability to develop NIH-funded research programs limited by inadequate or overly fragmented research start-up time and by inconsistent approaches to research mentoring. In concert, the recent abrupt decrease in rate of growth of NIH funding now threatens not only the pace of our progress, but survival of physician scientist activity within anesthesiology. Based on these observations, anesthesiology is at risk of losing its status as a respected academic discipline within the broader biomedical community. Based on historic analysis of periods of limited NIH budgets, critical analysis of anesthesiology research training programs relative to peer academic specialties,1 and considering the forecasted continued high demand for clinical manpower in our academic medical centers, we present the case for radical change in our approach to training within the specialty. Although effective solutions will require significant sacrifice by all sectors of the specialty, we propose that modest steps are far too late and will no longer succeed. This article, while seeking to inform and educate, is above all a passionate call for decisive action.

Considerable progress has been made in understanding the pathophysiology of perioperative stress responses and their impact on the cardiovascular system; however, researchers are just beginning to unravel genetic and molecular determinants that predispose to increased risk for postoperative cardiovascular adverse events. A new field, coined perioperative genomics, aims to apply functional genomic approaches to uncover the biological reasons why similar patients can have dramatically different clinical outcomes after surgery. For the perioperative physician, such findings may soon translate into prospective risk assessment incorporating genomic profiling of markers important in inflammatory, thrombotic, vascular, and neurologic responses to perioperative stress, with implications ranging from individualized additional pre-operative testing and physiological optimization, to perioperative decision-making, choice of monitoring strategies, and critical care resource utilization. We review current knowledge regarding genomic technologies in perioperative cardiovascular disease characterization and outcome prediction, as well as discuss future trends/challenges for translating integrated “omic” information into daily clinical management of the surgical patient.

Using an improved chromatin immunoprecipitation assay designed to increase immunoprecipitation efficiency, we investigated changes in RNA polymerase II (Pol II) density and carboxyl-terminal domain (CTD) phosphorylation during transcription of the cyclophilin A (PPIA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and several androgen-responsive genes in LNCaP cells. As generally observed in higher eukaryotes, promoter proximal pausing of Pol II appeared to occur on the PPIA and GAPDH genes, but apparently not on the androgen-responsive genes PSA and NKX3-1. Unlike some mammalian studies, we found that the CTD of Pol II in promoter regions contains little phosphorylation at Ser-2 of the heptad repeat, suggesting that Ser-2 phosphorylation is not involved in polymerase exit from the promoter region. In contrast, Pol II near the promoter displayed high levels of Ser-5 phosphorylation, which decreased as polymerase transcribed beyond the promoter region of the PPIA and GAPDH genes. However, total Pol II levels appear to decrease as much or more, suggesting that Ser-5 phosphorylation is maintained. In support of this conclusion, a phosphoserine 5-specific antibody quantitatively immunoprecipitates native hyperphosphorylated Pol II, suggesting that all polymerase with phosphoserine 2 also contains phosphoserine 5. Given reports indicating that phosphoserine 5 is present during elongation in yeast, our data suggest that gross changes in CTD phosphorylation patterns during transcription may be more conserved in yeast and humans than recognized previously.

We identified nine naturally-occurring human single nucleotide polymorphisms (SNPs) in the alpha(1a)-adrenoceptor (alpha(1a)AR) coding region, seven of which result in amino acid change. Utilizing rat-1 fibroblasts stably expressing wild type alpha(1a)AR or each SNP at both high and low levels, we investigated the effect of these SNPs on receptor function. Compared with wild type, two SNPs (R166K, V311I) cause a decrease in binding affinity for agonists norepinephrine, epinephrine, and phenylephrine, and also shift the dose-response curve for norepinephrine stimulation of inositol phosphate (IP) production to the right (reduced potency) without altering maximal IP activity. In addition, SNP V311I and I200S display altered antagonist binding. Interestingly, a receptor with SNP G247R (located in the third intracellular loop) displays increased maximal receptor IP activity and stimulates cell growth. The increased receptor signaling for alpha(1a)AR G247R is not mediated by altered ligand binding or a deficiency in agonist-mediated desensitization, but appears to be related to enhanced receptor-G protein coupling. In conclusion, four naturally-occurring human alpha(1a)AR SNPs induce altered receptor pharmacology and/or biological activity. This finding has potentially important implications in many areas of medicine and can be used to guide alpha(1a)AR SNP choice for future clinical studies.

α 1 -Adrenoceptors are G-protein-coupled receptors that bind catecholamines. Sixteen distinct human α 1A -adrenoceptor isoforms have been identified from human tissues, including five full-length and 11 truncated versions. An updated scheme for the identification of α 1A -adrenoceptor splice variants is proposed. Given the established roles of α 1 -adrenoceptors in benign prostatic hyperplasia, myocardial hypertrophy and other cardiovascular disorders, elucidation of the biological significance of the signaling diversity and potential pharmacological roles of α 1A -adrenoceptor splice variants are important areas of future research.

Alpha1a-adrenergic receptors (alpha1aARs) are present intracellularly and at the cell surface in cultured and natural cell models, where they are subject to agonist-mediated desensitization and internalization. To explore alpha1aAR trafficking, a hemagglutinin (HA)-tagged alpha1aAR/enhanced green fluorescent protein (EGFP) fusion protein was expressed in rat-1 fibroblasts and tracked by EGFP fluorescence and antibody labeling of surface receptors. Confocal analysis of antibody-labeled surface receptors revealed unexpected constitutive internalization in the absence of agonist stimulation. In partial agreement, the inverse agonist prazosin also caused a modest 20 +/- 2% increase in surface receptor levels, suggesting a partial block of constitutive internalization caused by decreased basal activation. However, prazosin was unable to prevent internalization of antibody-tagged surface receptors observed by confocal microscopy or cause obvious redistribution of intracellular receptor to the surface, suggesting that the alpha1aAR is internalizing even in a basal-inactive state. In contrast to the alpha1aAR, surface labeling of an HA-tagged alpha1b-EGFP fusion protein did not result in any apparent constitutive internalization. Constitutive internalization of the alpha1aAR seems to occur alongside reversible agonist-induced internalization, and both seem to involve clathrin-mediated endocytosis but not degradation in lysozymes. Surface receptor density must be maintained by recycling, because the protein synthesis inhibitor cycloheximide has no effect on total or surface receptor density in agonist-treated or untreated cells for 6 h. Constitutive agonist-independent trafficking of alpha1aARs may provide a novel mechanism by which an internal pool of alpha1aARs are maintained and recycled to allow continuous agonist-induced signaling.

Although α1AR antagonists have been used for more than two decades to treat lower urinary tract symptoms (LUTS), we have little understanding of the mechanistic basis of their efficacy and their role in the development of LUTS. It is clear that α1ARs play a critical role in bladder dysfunction and recent data suggest that α1AR subtype switching may play a key role in this pathophysiology, providing support for use of α1dAR-selective antagonists in treating irritative symptoms. This review seeks to summarize current levels of understanding in this field and discusses new concepts that suggest increased levels of complexity involving cross-talk in multiple receptor systems. Effective therapeutic modalities likely will involve increased subtype selective α1AR antagonists and other pharmacodynamic factors.

Recent studies reveal important and distinct roles for cardiac α1a adrenergic receptors (α1aARs). Surprisingly, given their importance in myocardial ischemia/reperfusion, hypoxia, and hypertrophy as well as frequent use of rat cardiomyocyte model systems, the rat α1aAR gene promoter has never been characterized. Therefore, we isolated 3.9 kb of rat α1aAR 5′-untranslated region and 5′-regulatory sequences and identified multiple transcription initiation sites. One proximal (P1) and several clustered upstream distal promoters (P2, P3, and P4) were delineated. Sequences surrounding both proximal and distal promoters lack typical TATA or CCAAT boxes but contain cis-elements for multiple myocardium-relevant nuclear regulators including Sp1, GATA, and CREB, findings consistent with enhanced cardiac basal α1aAR expression seen in Northern blots and reporter constructs. Promoter analysis using deletion reporter constructs reveals, in addition to a powerful upstream enhancer, a key region (–558/−542) important in regulating all α1aAR promoters with hypoxic stress. Gel shift analysis of this 14-bp region confirms a hypoxia-induced shift independent of direct hypoxia-inducible factor binding. Mutational analysis of this sequence identifies a novel 9-bp hypoxia response element, the loss of which severely attenuates hypoxia-mediated repression of α1aAR transcription. These findings for the α1a gene should facilitate elucidation of α1AR-mediated mechanisms involved in distinct myocardial pathologies.

Background This double-blind, randomized, controlled trial examined the effect of high-dose intrathecal bupivacaine in combination with general anesthesia on atrial beta-adrenergic receptor function, the stress response, and hemodynamics during coronary artery bypass graft surgery. Methods Thirty-eight patients were randomized to either control (n = 19) or intrathecal bupivacaine (ITB) groups (n = 19). Patients in the ITB group received 37.5 mg intrathecal hyperbaric bupivacaine before induction of general anesthesia. Control patients received an injection of local anesthetic into the skin and subcutaneous tissues (sham spinal). Comparisons were made between groups with respect to atrial receptor desensitization and down-regulation, in addition to circulating catecholamines and hemodynamics. Results In patients with cardiopulmonary bypass (CPB) times in excess of 1 h, the ITB group had significantly less atrial beta-receptor dysfunction, as measured by maximal isproteronol, 50% maximal isoproterenol, sodium fluoride-stimulated activity, and zinterol stimulation assays of adenylyl cyclase activity (P < or = 0.02) and beta-adrenergic receptor density (P = 0.02). Serum epinephrine, norepinephrine, and cortisol concentrations were significantly lower in the ITB group, independent of CPB times (P < 0.0001, P < 0.001, and P < 0.05, respectively). ITB patients had a higher cardiac index and a lower pulmonary vascular resistance index in the post-CPB time period (P < 0.01 and P < 0.05, respectively). In the pre-CPB period, mean arterial pressure and systemic vascular resistance index were significantly lower in the ITB group. Conclusions High-dose intrathecal bupivacaine, when combined with general anesthesia, resulted in less beta-receptor dysfunction and a lower stress response during coronary artery bypass graft surgery.

Transexamic acid (TXA) is an antifibrinolytic that has been used successfully to prevent blood loss during major surgery. However, as its usage has increased, there have been growing reports of postsurgical seizure events in cardiac surgery patients. In this issue of the JCI, Lecker et al. explore this connection and suggest that TXA-mediated inhibition of glycine receptors may underlie the effect. This finding prompted the authors to explore the preclinical efficacy of common anesthetics that function by reducing the TXA-mediated inhibition to prevent or modify postsurgical seizures.

Background Recent American Heart Association guidelines highlight the paucity of data on effectiveness and/or mechanisms underlying use of beta-adrenergic receptor (beta AR) antagonists after acute coronary syndromes in patients subsequently undergoing revascularization. It is important to assess whether beta AR antagonists might protect the heart and improve ventricular function in this scenario. The authors therefore used esmolol (an ultra-short-acting beta AR antagonist) to determine whether beta AR antagonist treatment improves left ventricular function in a canine model of acute reversible coronary ischemia followed by coronary reperfusion during cardiopulmonary bypass (CPB). The authors also tested whether the mechanism includes preserved beta AR signaling. Methods Dogs were randomized to either esmolol or saline infusions administered during CPB (n = 29). Pre-CPB and end-CPB transmyocardial left ventricular biopsies were obtained; plasma catecholamine concentrations, myocardial beta AR density, and adenylyl cyclase activity were measured. In addition, left ventricular systolic shortening and postsystolic shortening were determined immediately prior to each biopsy. Results While beta AR density remained unchanged in each group, isoproterenol-stimulated adenylyl cyclase activity decreased 26 +/- 6% in the control group but increased 38 +/- 10% in the esmolol group (pre-CPB to end-CPB, mean +/- SD, P = 0.0001). Left ventricular systolic shortening improved in both groups after release of coronary (LAD) ligature; however, the esmolol group increased to 72 +/- 23% of pre-CPB values compared to 48 +/- 12% for the control group (P = 0.0008). Conclusions These data provide prospective evidence that esmolol administration results in improved myocardial function. Furthermore, the mechanism appears to involve enhanced myocardial beta AR signaling.

Despite important roles in myocardial hypertrophy and benign prostatic hyperplasia, little is known about acute effects of agonist stimulation on α1a-adrenergic receptor (α1aAR) signaling and function. Regulatory mechanisms are likely complex since 12 distinct human α1aAR carboxyl-terminal splice variants have been isolated. After determining the predominance of the α1a-1AR isoform in human heart and prostate, we stably expressed an epitope-tagged α1a-1AR cDNA in rat-1 fibroblasts and subsequently examined regulation of signaling, phosphorylation, and internalization of the receptor. Human α1aAR-mediated inositol phosphate signaling is acutely desensitized in response to both agonist and phorbol 12-myristate 13-acetate (PMA) exposure. Concurrent with desensitization, α1aARs in32Pi-labeled cells are rapidly phosphorylated in response to both NE and PMA stimulation. Despite the ability of PKC to desensitize α1aARs when directly activated with PMA, inhibitors of PKC have no effect on agonist-mediated desensitization. In contrast, involvement of GRK kinases is suggested by the ability of GRK2 to desensitize α1aARs. Internalization of cell surface α1aARs also occurs in response to agonist stimulation (but not PKC activation), but is initiated more slowly than receptor desensitization. Significantly, deletion of the α1aAR carboxyl terminus has no effect on receptor internalization or either agonist-induced or GRK-mediated receptor desensitization. Because mechanisms underlying acute agonist-mediated regulation of human α1aARs are primarily independent of the carboxyl terminus, they may be common to all functional α1aAR isoforms.

Purpose: α1-Adrenergic receptor (α1AR) antagonists are effective for relieving obstructive and irritative symptoms in patients with bladder outlet obstruction. While the α1aAR is responsible for prostate smooth muscle relaxation and outlet obstruction relief, to our knowledge the mechanisms underlying the relief of irritative symptoms remain to be determined. Therefore, we investigated mechanisms by which bladder α1AR subtypes may be involved in this process.Materials and Methods: We studied 42 rats, including 6 unoperated controls, 17 sham operated controls and 19 obstructed animals. Animals were characterized for baseline voiding pattern, followed by surgical intervention or sham surgery to establish obstruction (1.09 mm. restricted opening). After 6 weeks to enable the development of detrusor hypertrophy, voiding behavior was reexamined, the animals were sacrificed and bladder tissue was immediately placed in liquid nitrogen. α1AR subtype messenger (m)RNA was quantitated using quantitative competitive ...

Objectives: Alpha1-Adrenergic Receptors (α1ARs), members of the G protein-coupled receptor (GPCR) superfamily, play a major role in regulating cardiovascular (CV) function. Recently, we discovered that naturally occurring human α1aAR-G247R (247R) genetic variant, identified in the 3rd intracellular loop (3iL) of the receptor in highly hypertensive patient, triggers constitutive hyperproliferation in fibroblasts, cardiomyoblasts and smooth muscle cells (SMC). Specific proteins mediating this signaling remain unknown. Spinophilin (SPL) is a ubiquitously expressed protein controlling GPCR signaling by binding its 3rd intracellular loop (3iL). We hypothesize that SPL mediates α1aAR signaling and examined whether SPL directly interacts with α1aAR-WT (WT) and 247R. Methods: Cells were co-transfected with HA-α1ARs and full length Myc-SPL or its fragments to determine SPL domains responsible for binding to α1ARs. Cell lysates were co-immunoprecipitated with HA-tag antibodies. SPL levels were analyzed by Western blotting. SPL knockdown experiments were performed by transiently transfecting cells with SPL or scrambled siRNA, cell proliferation was determined by cell counting. Results: We demonstrate a distinct interaction of SPL with WT and 247R, WT interaction being the strongest. Different domains of SPL differentially interact with WT or 247R. SPL 1-480aa fragment interacts stronger with WT indicating interaction with 3iL, while SPL 480-817 fragment interacts stronger with 247R. Endogenous SPL levels are increased in 247R cells compared to WT or control cells. Inhibition of SPL expression in SMC with siRNA reduces 247R-triggered hyperproliferation to near normal levels and has no effect in WT cells. Conclusions: We identified SPL as a novel interacting protein involved in mediating intracellular signaling of α1aAR and its genetic variant in CV cells. Different domains of SPL differentially bind to WT or 247R indicating that SPL has a distinct role in regulation of their signaling pathways. Our findings also reveal that SPL is critical for 247R-triggered EGFR transactivation pathways. Thus SPL could be considered as a potentially novel target in α1aAR-mediated cardiovascular disorders.

Activation of α 1 -Adrenergic Receptors (α 1 ARs), members of the G protein-coupled receptor (GPCR) superfamily, in response to stimulation of the sympathetic nervous system by catecholamines plays a major role in regulating cardiovascular (CV) function. Among three α 1 AR subtypes (α 1a ,α 1b ,α 1d ), α 1a ARs predominate in human resistant vessels and in heart. Recently, we discovered that naturally occurring human α 1a AR-G247R (247R) genetic variant, identified in the 3 rd intracellular loop (3iL) of the receptor in highly hypertensive patient, triggers constitutive hyperproliferation in CV cells (cardiomyoblasts, smooth muscle cells (SMC) and fibroblasts), which may lead to myocardial fibrosis and remodeling. In fibroblasts and cardiomyoblasts 247R triggered hyperproliferation is due to constitutive active coupling to Gq-independent βarrestin1/MMP/EGFR/ERK dependent pathway, while in SMC it is Gq- and MMP/EGFR/ERK-dependent. Here we report that α 1a AR-WT (WT) and 247R differentially interact with ubiquitous multi-domain scaffold protein spinophilin (SPL) that binds to 3iL of several GPCRs competing with arrestin thereby prolonging their signaling. The role of SPL in CV regulation is poorly studied. We hypothesized that SPL mediates constitutive signaling of 247R and examined whether SPL directly interacts with α 1a AR-WT or 247R. Our preliminary results reveal a direct interaction of SPL with WT and 247R: the SPL-WT interaction appears to be stronger as determined by co-immunoprecipitation. Different domains of SPL differentially interact with WT or 247R. SPL 1-480aa fragment interacts stronger with WT indicating interaction with 3iL, while SPL 480-817 fragment interacts stronger with 247R. Our preliminary results also demonstrate that 247R expression in all three cell types elevates endogenous SPL protein levels. Importantly, inhibition of SPL expression with specific siRNA reduces 247R-triggered hyperproliferation in SMC and cardiomyoblasts to near normal levels, while SPL knockdown has no effect in WT cells. Thus, we identified SPL as a novel protein involved in interacting and signaling of α 1a AR and its genetic variant in CV cells and that SPL could be considered as a potentially novel target in α 1a AR-mediated cardiovascular disorders.

Activation of α1-Adrenergic Receptors (α1ARs), members of the G protein-coupled receptor (GPCR) superfamily, in response to stimulation of the sympathetic nervous system by catecholamines plays a major role in regulating cardiovascular (CV) function and blood pressure (BP). Among three α1AR subtypes (α1a,α1b,α1d) expressed in vasculature α1aARs predominate in human vascular smooth muscle cells (SMC), particularly in resistant vessels most involved in BP control. Polymorphisms in the α1aAR gene are also related to hypertension (HTN), one of the major CV risk factors. Recently, we discovered a novel, unique mechanism of hypertension triggered by naturally occurring human α1aAR-G247R (247R) genetic variant, identified in the 3rd intracellular loop (3iL) of the receptor in hypertensive patient. In fibroblasts, 247R signals via constitutive active coupling to the βarrestin1/MMP7/EGFR pathway. Here we report that 247R expression in SMC triggers constitutive, Gq- and MMP/EGFR/ERK-dependent hyperproliferation. Agonist (phenylephrine) treatment of cells inhibits hyperproliferation and induces hypertrophy and α1aAR inverse agonist prazosin inhibits hyperproliferation and hypertrophy indicating Gq-dependent pathway. Expression of 247R in SMC also triggers upregulation of spinophilin (SPL), a ubiquitous multi-domain scaffold protein that binds to 3iL of several GPCRs and competes with βarrestins for 3iL binding. We hypothesized that SPL mediates constitutive signaling of 247R and examined whether SPL directly interacts with α1aAR-WT or 247R. Our preliminary results demonstrate a distinct interaction of α1aAR-WT and 247R with SPL, although the SPL-α1AR-WT interaction appears to be strongest. SPL 1-480aa fragment demonstrates stronger interaction with α1aAR-WT indicating that this domain is responsible for interaction with 3iL. Interestingly the SPL 480-817 fragment has stronger interaction with 247R. Thus, expression of the naturally occurring human α1aAR genetic variant in vascular cells activates distinct signaling pathways leading to aberrant hyperproliferation and hypertrophy, and may eventually result in hypertension and other CV diseases suggesting a possible novel mechanism underlying some forms of human hypertension.

In most investigations, genetic and environmental factors have been shown to interact in altering the progression of ageing related disease including stroke and cognitive decline. The peri-operative period appears to be no different, with environmental (operative) and genetic factors interacting to determine the presence, absence or severity of neurological outcomes. In our initial work we confirmed that apolipoprotein E4 (apoE4) alters cognitive function after coronary artery bypass graft (CABG), the mechanism of this association apparently being related to atherosclerosis and inflammatory factors. These findings are consistent with multiple studies that have shown an important role of apoE in the modulation of neurological injury and recovery following a variety of acute ischaemic insults including intra-cerebral haemorrhage, closed head injury, acute stroke and dementia pugilistica. More recently we have found that polymorphism of the GPIIIa constituent of the platelet integrin receptor, GPIIb/IIIa (PL A2 ) polymorphism is also associated with early post-operative cognitive decline. The association of multiple genetic factors with the incidence or severity of neurological injury is not surprising, since the factors that alter cerebral injury and recovery are complex. Similarly, there are genetic factors that determine the occurrence of tissue injury and its recovery, including genes that modulate atherosclerosis, emboli, inflammation, thrombosis and vascular reactivity. We have only begun to scratch the surface in our understanding of functional genomics as it applies to peri-operative outcomes. Functional genomics, as applied to complex human disease, will require a unique combination of cutting-edge genetics and large, highly phenotyped patient populations in order to further our understanding of the complex genetic–environmental interactions. This chapter will concentrate initially on the broad classes of genetic factors that may play a role in neurological injury and recovery, and will conclude with our current data on those genetic factors that predict neurocognitive decline after cardiac surgery and the potential mechanisms driving these associations.

Purpose: We established an immortalized human prostate stromal cell line with retained markers of cell differentiation and α1-adrenergic receptor expression.Materials and Methods: Primary human prostate stromal explants were infected with an amphotrophic retrovirus encoding the E6/E7 open reading frame of the human papillomavirus type 16. Immunohistochemistry was used to verify the expression of prostate stromal markers. α1-Adrenergic receptor expression was investigated using ribonuclease protection assays and radioligand binding. Cell proliferation was measured by the WST-1 † assay and cell counting.Results: Clonal isolates of individual prostate stromal cells were isolated and passed in selection media. E6 and E7 expression was verified using reverse transcriptase polymerase chain reaction in the selected cell line. The new prostate stromal cell line PS30 was established which maintains the expression of α-smooth muscle actin and expresses 22 fmol./mg. of protein of α1-adrenergic receptors, approximate...

Adrenergic receptors (ARs) are members of the G-protein-coupled receptor family, which includes alpha 1ARs, alpha 2ARs, beta 1ARs, beta 2ARs, beta 3ARs, adenosine, muscarinic, angiotensin, endothelin receptors, and many others that are responsible for a large variety of physiologic effects through G-protein coupling. This review focuses on alpha 1ARs and their regulation at both the mRNA and protein levels. Currently, three alpha 1AR subtypes have been characterized both pharmacologically and at the gene level: alpha 1aAR, alpha 1bAR, and alpha 1dAR. These are expressed in a species- and tissue-dependent manner. Mutagenesis approaches have been extremely valuable in the identification of key residues that govern alpha 1AR ligand binding and signaling. These studies reveal that alpha 1ARs have evolved an exquisitely sensitive regulation of their activity in which any disruption of the native structure has profound effects on subsequent function and effector coupling. Significant advances have also been made in the elucidation of signaling pathway components, resulting in the identification of novel pathways that can lead to pathologic conditions. Specific topics include mitogen-activated protein kinase, phosphatidylinositol 3-kinase, and G-protein-coupled receptor cross-talk pathways. Within this context, recent studies identifying underlying transcriptional mechanisms involved in the regulation of the alpha 1AR subtypes are also discussed. Finally, given the potentially important role of alpha 1ARs in the vasculature, as well as in the pathology of many diseases, such as myocardial hypertrophy and benign prostatic hyperplasia, the clinical relevance of alpha 1AR distribution, pharmacology, and therapeutic intervention is reviewed.

Abstract —Cardiac G protein–coupled receptors that couple to Gα s and stimulate cAMP formation (eg, β-adrenergic, histamine, serotonin, and glucagon receptors) play a key role in cardiac inotropy. Recent studies in rodent cardiac myocytes and transfected cells have revealed that one of these receptors, the β 2 -adrenergic receptor (AR), also couples to the inhibitory G protein Gα i (activation of which inhibits cAMP formation). If β 2 ARs could be shown to couple to Gα i in the human heart, it would have important ramifications, because levels of Gα i increase with age and in failing human heart. Therefore, we investigated whether β 2 ARs in the human heart activate Gα i . By photoaffinity labeling human atrial membranes with [ 32 P]azidoanilido-GTP, followed by immunoprecipitation with antibodies specific for Gα i , we found that Gα i is activated by stimulation of β 2 ARs but not of β 1 ARs. In addition, we found that other Gα s -coupled receptors also couple to Gα i , including histamine, serotonin, and glucagon. When coupling of these receptors to Gα i is disrupted by pertussis toxin, their ability to stimulate adenylyl cyclase is enhanced. These data provide the first evidence that β 2 AR and many other Gα s -coupled receptors in human atrium also couple to Gα i and that abolishing the coupling of these receptors to Gα i increases the receptor-mediated adenylyl cyclase activity.

Background Nonselective alpha2-adrenergic receptor (alpha2AR) agonists (e.g., clonidine) mediate antinociception in part through alpha2ARs in spinal cord dorsal horn; however, use of these agents for analgesia in humans is limited by unwanted sedation and hypotension. The authors previously demonstrated alpha2a approximately alpha2b > > > alpha2c mRNA in human spinal cord dorsal horn cell bodies. However, because 20% of dorsal horn alpha2ARs derive from cell bodies that reside in the associated dorsal root ganglion (DRG), it is important to evaluate alpha2AR expression in this tissue as well. Therefore, the authors evaluated the hypothesis that alpha2b mRNA, alpha2c mRNA, or both are present in human DRG. Methods Molecular approaches were used to determine alpha2AR expression in 28 human DRGs because of low overall receptor mRNA expression and small sample size. After creation of synthetic competitor cDNA and establishment of amplification conditions with parallel efficiencies, competitive reverse transcription polymerase chain reaction was performed using RNA isolated from human DRG. Results Overall expression of alpha2AR mRNA in DRG is low but reproducible at all spinal levels. alpha2b and alpha2cAR subtype mRNAs predominate (alpha2b approximately alpha2c), accounting for more than 95% of the total alpha2AR mRNA in DRG at all human spinal nerve root levels. Conclusions Predominance of alpha2b and alpha2cAR mRNA in human DRG is distinct from alpha2AR mRNA expression in cell bodies originating in human spinal cord dorsal horn, where alpha2a and alpha2b predominate with little or absent alpha2c expression. These findings also highlight species heterogeneity in alpha2AR expression in DRG. If confirmed at a protein level, these findings provide an additional step in unraveling mechanisms involved in complex neural pathways such as those for pain.

Purpose: To investigate the level of expression, activation state, and functional significance of extracellular signal regulated kinase (ERK) in prostate cancer.Materials and Methods: Human prostate tissue samples (n = 22) were obtained from patients undergoing radical prostatectomy for localized adenocarcinoma of the prostate (n = 16, age range 44 to 72 years) or normal prostate specimens (n = 6, age ranges 19 to 47 years) obtained from rapid autopsy. Immunoblots, in vitro kinase assays, and immunohistochemistry were used to determine the expression and activation state of ERK in human prostate cancer.Results: Immunoblot and in vitro kinase assays demonstrated a 15-fold increase in ERK activation in prostate cancer specimens compared with normal human prostate tissue; however, ERK expression levels were only 1.3-fold higher in cancer. Immunohistochemical analysis demonstrated similar expression of ERK in cancer and normal tissues; however, phosphorylated ERK demonstrated greater intensity in the ...

α1-Adrenergic receptors (α1ARs) are G protein-coupled receptors important in the dynamic component of benign prostatic hyperplasia. α1ARs stimulate predominantly phosp