Defeating diabetes in the desert: A community-based mHealth diabetes screening intervention in Jodhpur Rajasthan.

Background: There is a paucity of demonstrated models for mHealth-based diabetes screening and coordinated care in India, especially in western Rajasthan, which is the part of Thar desert. Materials and Methods: JSPH collaboratively developed and implemented an easy-to-use, noninvasive, mobile phone-based screening interview, to identify adults at high risk for diabetes. The high risk for diabetes was defined using multiple clinical and epidemiologic criteria, all based on the evidence for India and globally. Since participants above 35 years or older were only considered in the screening, the application was designed to categorize the participants as high and low risk. Results: Out of 4000 screened participants, the percentage of males and females were 51% and 50%, respectively. Participants found to be at high risk and low risk were n = 3600 (90%) and 400 (10%). The mean age of high- and low-risk participants was 52.2 (+12.8) and 36.2 (+4.2), respectively. Of the 3600 high-risk individuals who have been given a follow-up interview, 90.50% of high-risk individuals obtained diabetes testing, and of these, 65.67% had a written report showing they test positive for diabetes or prediabetes, requiring ongoing clinical care. Conclusions: JSPH mHealth application provided a novel noninvasive way to better identify those at high diabetes risk in the community and demonstrated how to optimize the use of mobile health methods in diabetes prevention and care services.

Burden of sickle cell disease in tribal students in Maa-Baadi institutions in southern Rajasthan - A pilot study.

Background & objectives: Sickle cell disease (SCD), an inherited disorder of erythrocytes, is highly prevalent in the tribal population of India. The tribal population of India is approximately 100 million and it is necessary to identify the magnitude of this problem. Furthermore, the prevalence of the disease is unknown among the five million tribal people of southern provinces of Rajasthan. In this study, we intended to determine the prevalence and characteristics of sickle cell disorder among the tribal inhabitants of southern Rajasthan. Methods: This cross-sectional study was conducted among the tribal students of the Maa-Baadis and hostels situated in the five tribal sub-plan districts of Rajasthan. Maa-Baadi centres are located in every village, whereas for every four to five villages, one hostel is allocated to accommodate the tribal students. The screening for SCD was done by solubility test and electrophoresis was used for confirmation. Results: A total of 36,752 tribal students were screened from 1,006 Maa-Baadi centres and 243 hostels. The prevalence of SCD among the tribal students was 5.8 per cent. The prevalence of heterozygous and homozygous conditions was 5.61 and 0.17 per cent, respectively. Among the five sub-plan districts, the highest prevalence was observed in Sirohi district (10.5%) followed by Banswara (7.42%), Udaipur (6.53%), Pratapgarh (5.51%) and Dungarpur (1.89%). Among the four major tribes belonging to these districts, the highest prevalence was recorded in Garasia tribes (13.81%). The history of leg ulcers and the mean pulse rate were significantly high in SCD individuals. Interpretation & conclusions: SCD is a significant problem among the tribes of southern Rajasthan, with the highest prevalence among the Garasia tribe. The present study recommends that a structured screening programme targeting the entire tribal population with appropriate counselling as well as providing treatment through the existing health system is the need of the hour.

Effectiveness and feasibility of methanol extracted latex of Calotropis procera as larvicide against dengue vectors of western Rajasthan, India.

BACKGROUND & OBJECTIVES: Identification of novel effective larvicide from natural resources is essential to combat developing resistances, environmental concerns, residue problems and high cost of synthetic insecticides. Results of earlier laboratory findings have shown that Calotropis procera extracts showed larvicidal, ovicidal and refractory properties towards ovipositioning of dengue vectors; further, latex extracted with methanol was found to be more effective compared to crude latex. For testing efficacy and feasibility of extracted latex in field, the present study was undertaken in different settings of Jodhpur City, India against dengue vectors. METHODS: Study areas were selected based on surveillance design for the control of dengue vectors. During the study period domestic and peri-domestic breeding containers were treated with methanol extracted latex and mortality was observed after 24 h as per WHO guidelines. Latex was manually collected from internodes of Calotropis procera and extracted using methanol (AR) grade. RESULTS: Methanol extracted latex of C. procera was found effective and feasible larvicide against dengue vectors in the field conditions. Cement tanks, clay pots and coolers (breeding sites) were observed as key containers for the control of dengue transmission. INTERPRETATION & CONCLUSION: Today environmental safety is considered to be very important. Herbal composition prepared by the extraction of latex of C. procera can be used as an alternative approach for the control of dengue vectors. This will reduce the dependence on expensive products and stimulate local efforts to enhance the public involvement.

Oxidized Ca(2+)/calmodulin-dependent protein kinase II triggers atrial fibrillation.

BACKGROUND: Atrial fibrillation (AF) is a growing public health problem without adequate therapies. Angiotensin II and reactive oxygen species are validated risk factors for AF in patients, but the molecular pathways connecting reactive oxygen species and AF are unknown. The Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) has recently emerged as a reactive oxygen species-activated proarrhythmic signal, so we hypothesized that oxidized CaMKIIδ could contribute to AF. METHODS AND RESULTS: We found that oxidized CaMKII was increased in atria from AF patients compared with patients in sinus rhythm and from mice infused with angiotensin II compared with mice infused with saline. Angiotensin II-treated mice had increased susceptibility to AF compared with saline-treated wild-type mice, establishing angiotensin II as a risk factor for AF in mice. Knock-in mice lacking critical oxidation sites in CaMKIIδ (MM-VV) and mice with myocardium-restricted transgenic overexpression of methionine sulfoxide reductase A, an enzyme that reduces oxidized CaMKII, were resistant to AF induction after angiotensin II infusion. CONCLUSIONS: Our studies suggest that CaMKII is a molecular signal that couples increased reactive oxygen species with AF and that therapeutic strategies to decrease oxidized CaMKII may prevent or reduce AF.

Genetic inhibition of Na+-Ca2+ exchanger current disables fight or flight sinoatrial node activity without affecting resting heart rate.

RATIONALE: The sodium-calcium exchanger 1 (NCX1) is predominantly expressed in the heart and is implicated in controlling automaticity in isolated sinoatrial node (SAN) pacemaker cells, but the potential role of NCX1 in determining heart rate in vivo is unknown. OBJECTIVE: To determine the role of Ncx1 in heart rate. METHODS AND RESULTS: We used global myocardial and SAN-targeted conditional Ncx1 knockout (Ncx1(-/-)) mice to measure the effect of the NCX current on pacemaking activity in vivo, ex vivo, and in isolated SAN cells. We induced conditional Ncx1(-/-) using a Cre/loxP system. Unexpectedly, in vivo and ex vivo hearts and isolated SAN cells showed that basal rates in Ncx1(-/-) (retaining ≈20% of control level NCX current) and control mice were similar, suggesting that physiological NCX1 expression is not required for determining resting heart rate. However, increases in heart rate and SAN cell automaticity in response to isoproterenol or the dihydropyridine Ca(2+) channel agonist BayK8644 were significantly blunted or eliminated in Ncx1(-/-) mice, indicating that NCX1 is important for fight or flight heart rate responses. In contrast, the pacemaker current and L-type Ca(2+) currents were equivalent in control and Ncx1(-/-) SAN cells under resting and isoproterenol-stimulated conditions. Ivabradine, a pacemaker current antagonist with clinical efficacy, reduced basal SAN cell automaticity similarly in control and Ncx1(-/-) mice. However, ivabradine decreased automaticity in SAN cells isolated from Ncx1(-/-) mice more effectively than in control SAN cells after isoproterenol, suggesting that the importance of NCX current in fight or flight rate increases is enhanced after pacemaker current inhibition. CONCLUSIONS: Physiological Ncx1 expression is required for increasing sinus rates in vivo, ex vivo, and in isolated SAN cells, but not for maintaining resting heart rate.

Calmodulin-dependent protein kinase II: linking heart failure and arrhythmias.

Understanding relationships between heart failure and arrhythmias, important causes of suffering and sudden death, remains an unmet goal for biomedical researchers and physicians. Evidence assembled over the past decade supports a view that activation of the multifunctional Ca(2+) and calmodulin-dependent protein kinase II (CaMKII) favors myocardial dysfunction and cell membrane electrical instability. CaMKII activation follows increases in intracellular Ca(2+) or oxidation, upstream signals with the capacity to transition CaMKII into a Ca(2+) and calmodulin-independent constitutively active enzyme. Constitutively active CaMKII appears poised to participate in disease pathways by catalyzing the phosphorylation of classes of protein targets important for excitation-contraction coupling and cell survival, including ion channels and Ca(2+) homeostatic proteins, and transcription factors that drive hypertrophic and inflammatory gene expression. This rich diversity of downstream targets helps to explain the potential for CaMKII to simultaneously affect mechanical and electrical properties of heart muscle cells. Proof-of-concept studies from a growing number of investigators show that CaMKII inhibition is beneficial for improving myocardial performance and for reducing arrhythmias. We review the molecular physiology of CaMKII and discuss CaMKII actions at key cellular targets and results of animal models of myocardial hypertrophy, dysfunction, and arrhythmias that suggest CaMKII inhibition may benefit myocardial function while reducing arrhythmias.

Oxidized CaMKII causes cardiac sinus node dysfunction in mice.

Sinus node dysfunction (SND) is a major public health problem that is associated with sudden cardiac death and requires surgical implantation of artificial pacemakers. However, little is known about the molecular and cellular mechanisms that cause SND. Most SND occurs in the setting of heart failure and hypertension, conditions that are marked by elevated circulating angiotensin II (Ang II) and increased oxidant stress. Here, we show that oxidized calmodulin kinase II (ox-CaMKII) is a biomarker for SND in patients and dogs and a disease determinant in mice. In wild-type mice, Ang II infusion caused sinoatrial nodal (SAN) cell oxidation by activating NADPH oxidase, leading to increased ox-CaMKII, SAN cell apoptosis, and SND. p47-/- mice lacking functional NADPH oxidase and mice with myocardial or SAN-targeted CaMKII inhibition were highly resistant to SAN apoptosis and SND, suggesting that ox-CaMKII-triggered SAN cell death contributed to SND. We developed a computational model of the sinoatrial node that showed that a loss of SAN cells below a critical threshold caused SND by preventing normal impulse formation and propagation. These data provide novel molecular and mechanistic information to understand SND and suggest that targeted CaMKII inhibition may be useful for preventing SND in high-risk patients.

Proarrhythmic defects in Timothy syndrome require calmodulin kinase II.

BACKGROUND: Timothy syndrome (TS) is a disease of excessive cellular Ca(2+) entry and life-threatening arrhythmias caused by a mutation in the primary cardiac L-type Ca(2+) channel (Ca(V)1.2). The TS mutation causes loss of normal voltage-dependent inactivation of Ca(V)1.2 current (I(Ca)). During cellular Ca(2+) overload, the calmodulin-dependent protein kinase II (CaMKII) causes arrhythmias. We hypothesized that CaMKII is a part of the proarrhythmic mechanism in TS. METHODS AND RESULTS: We developed an adult rat ventricular myocyte model of TS (G406R) by lentivirus-mediated transfer of wild-type and TS Ca(V)1.2. The exogenous Ca(V)1.2 contained a mutation (T1066Y) conferring dihydropyridine resistance, so we could silence endogenous Ca(V)1.2 with nifedipine and maintain peak I(Ca) at control levels in infected cells. TS Ca(V)1.2-infected ventricular myocytes exhibited the signature voltage-dependent inactivation loss under Ca(2+) buffering conditions, not permissive for CaMKII activation. In physiological Ca(2+) solutions, TS Ca(V)1.2-expressing ventricular myocytes exhibited increased CaMKII activity and a proarrhythmic phenotype that included action potential prolongation, increased I(Ca) facilitation, and afterdepolarizations. Intracellular dialysis of a CaMKII inhibitory peptide, but not a control peptide, reversed increases in I(Ca) facilitation, normalized the action potential, and prevented afterdepolarizations. We developed a revised mathematical model that accounts for CaMKII-dependent and CaMKII-independent effects of the TS mutation. CONCLUSIONS: In TS, the loss of voltage-dependent inactivation is an upstream initiating event for arrhythmia phenotypes that are ultimately dependent on CaMKII activation.

Entomological studies for surveillance and prevention of dengue in arid and semi-arid districts of Rajasthan, India.

BACKGROUND & OBJECTIVES: Rajasthan is one of the dengue endemic states of India. Very few studies have been published on entomological aspects of dengue in this state. Owing to water scarcity, inhabitants in desert areas overstore domestic water which leads to the persistence of dengue vectors within the domestic premises. Area specific knowledge on breeding, key containers and seasonal rhythms of vector population is essential for preparing an effective prevention plan against dengue. Present paper reports results of entomological investigations on dengue vectors in arid and semi-arid districts of Rajasthan. METHODS: Longitudinal studies were undertaken during 2004-06 in one arid and two semi-arid dengue endemic districts of Rajasthan. Adult and larval Aedes were collected from the randomly selected houses in representative towns and villages with associated details of container types and water storage practices of inhabitants. RESULTS: In urban areas during all the seasons adult house index (AHI) of Aedes aegypti was maximum in desert zone (25) and least in semi-arid area with saline river III (1). The difference of AHI during three seasons was statistically significant (chi2 = 16.1, p < 0.01 for urban; and chi2 = 50.71, p < 0.001 for rural). Breeding of Ae. aegypti among urban settings was maximum in desert zone. During all the seasons cement tanks were the key breeding habitats for Ae. aegypti in desert as well as semi-arid areas. INTERPRETATION & CONCLUSION: Water storage habits during summer season emerged to be the risk factor of vector abundance in urban areas of arid and semi-arid settings. A carefully designed study of key containers targeting cement tanks as the primary habitats of mosquito control may lead to commendable results for dengue prevention.

Importance of socioeconomic status and tree holes in distribution of Aedes mosquitoes (Diptera: Culicidae) in Jodhpur, Rajasthan, India.

Immature Aedes mosquitoes were found in domestic, peridomestic, and tree hole habitats within three socioeconomic strata of Jodhpur, a city within an arid area of Rajasthan, India, endemic for dengue. Peridomestic habitats served as a persistent source of Aedes vectors, especially those used for watering cows for religious reasons that were located within high socioeconomic areas. Domestic (indoor) containers within low socioeconomic strata showed a higher container index (27.0%) than periurban areas with cattle sheds (14.3%) or high socioeconomic areas (18.1%). Mosquitoes were collected in tree holes at zoos and gardens supporting several species of monkeys. Six of 67 Aedes albopictus Skuse reared from immatures collected in tree holes tested positive for dengue antigen acquired through vertical transmission, possibly indicating a persistence mechanism for dengue virus within an urban environment.

Stable expression of constitutively activated mutant h5HT6 and h5HT7 serotonin receptors: inverse agonist activity of antipsychotic drugs.

RATIONALE: In order to determine the possible relationship between antipsychotic drug properties and inverse agonist activity at h5HT6 and h5HT7 receptors, constitutively activated forms of these receptors were created by site-specific mutagenesis. Typical and atypical antipsychotic drugs were assayed for their potencies as inverse agonists at these mutated receptors. OBJECTIVES: Stable cell lines expressing constitutively activated forms of the h5HT6 and h5HT7 receptors were created. Typical and atypical antipsychotic drugs demonstrating high to moderate affinities for the h5HT6 and h5HT7 receptors were assayed for their potencies in reversing the agonist-independent activity (inverse agonist activity). RESULTS: The E322R h5HT6 mutant and the S267K h5HT7 mutant displayed sufficiently robust agonist-independent activity when expressed in stable cell lines to allow the detailed, concentration-dependent, investigation of the inverse agonist activity of typical and atypical antipsychotic drugs. All the drugs tested displayed inverse agonist activity at both the activated h5HT6 and h5HT7 receptors. Native forms of these receptors did not display any constitutive activity. Interestingly, atypical antipsychotic drugs displayed potent inverse agonist activity, relative to typical antipsychotic drugs, at the h5HT7 receptor. LSD displayed neutral antagonist properties at the mutant h5HT7 receptor. CONCLUSIONS: Site-specific mutations in the third intracellular loop of the G(s)-coupled h5HT6 and h5HT7 receptors produce constitutive activation. Antipsychotic drugs display inverse agonist activity at the activated receptors. The inverse agonist mechanism-of-action of the atypical antipsychotic drugs at the h5HT7 receptors may be different from the typical antipsychotic drugs as these drugs displayed far higher potencies as inverse agonists at the h5HT7 receptor.

Arylguanidine and arylbiguanide binding at 5-HT3 serotonin receptors: a QSAR study.

For a series of monosubstituted arylguanidines, 5-HT3 receptor affinity was found generally related to the electron withdrawing nature of the substituent at the aryl 3-position and the lipophilicity of the 4-position substituent. A broader examination of 35 arylguanidines and arylbiguanides revealed that affinity could be described by molecular polarizability, a Chi index term (8chiP), and the sum of all (-Cl) E-State values (SsCl) in the molecule.

Binding of tetrahydrocarboline derivatives at human 5-HT5A receptors.

On the basis of an earlier finding that 5-methyl-5H-1,2,3,4-tetrahydropyrido[4,3-b]indole (5-methyl-1,2,3,4-tetrahydro-gamma-carboline; 1) binds at murine 5-HT(5A) receptors, preliminary structure-affinity studies were conducted. The present investigation extends these structure-affinity studies using human 5-HT(5A) receptors and examined additional analogues of 1. It was found (a) that there is little interspecies difference for the affinities of these compounds, (b) that an intact 1,2,3,4-tetrahydro-gamma-carboline ring system seems optimal and an N(2)-(3-(substituted-phenoxy)propyl) moiety results in high affinity, (c) that structurally related 1,2,3,4-tetrahydro-beta-carbolines also bind at 5-HT(5A) receptors, and (d) that all examined derivatives also possess affinity for 5-HT(2A) receptors. Evidence is provided that 5-HT(5A) and 5-HT(2A) receptor affinities probably do not covary and that it might be possible, with continued investigation, to develop analogues with enhanced 5-HT(5A) selectivity.

Conformationally-restricted analogues and partition coefficients of the 5-HT3 serotonin receptor ligands meta-chlorophenylbiguanide (mCPBG) and meta-chlorophenylguanidine (mCPG).

The present investigation examined two features of arylbiguanide and arylguanidine 5-HT(3) ligands: conformation and partition coefficients. Several conformationally-constrained analogues of mCPBG (2) and mCPG (11; K(i)=32 nM) were prepared and of these only 2-amino-5-chloro-3,4-dihydroquinazoline (14; K(i)=34 nM) retained high affinity. The partition coefficient of compound 11 (LogP(app)=-0.64) was less than that of its corresponding arylbiguanide 2 (LogP(app)=-0.38). The quinazoline structure may represent a pharmacologically-active conformation of these agents, and the arylbiguanides were found more lipid soluble than their arylguanidine counterparts at physiological pH.

gamma-Carbolines: binding at 5-HT5A serotonin receptors.

Screening of various agents resulted in the identification of 5-methyl-1,2,3,4-tetrahydro-gamma-carboline (1; K(i)=5,300 nM) as a compound with modest affinity for mouse 5-HT(5A) receptors. Structure-affinity studies were conducted resulting in 5-methyl-2-[3-(4-fluorophenoxy)propyl]-1,2,3,4-tetrahydro-gamma-carboline (17; K(i)=13 nM). Although 17 also binds at 5-HT(2) receptors, it serves as a novel lead for the further development of 5-HT(5A) ligands.

Creation, expression, and characterization of a constitutively active mutant of the human serotonin 5-HT6 receptor.

The serotonin 5-HT(6) receptor, a G-protein-coupled receptor, displays high affinity for antipsychotic, antidepressant, and psychotropic drugs. We created a constitutively active form of the human 5-HT(6) receptor in order to probe the molecular domains of receptor activation and to determine if inverse agonist activities of antipsychotic drugs contribute to their clinical profile. Previous studies from our laboratory support a critical role for the c-terminal region of the third intracellular loop (il3) in the activation of G(q)-coupled serotonin receptors. In the present study, PCR-based mutagenesis was used to mutate serine 267 (S6.34) in the c-terminal region of il3 to lysine (S267K). The native and S267K 5-HT(6) receptors were expressed in COS-7 cells to study the functional effects of the mutation. The S267K receptor shows 10-fold higher affinity for serotonin than the native receptor and demonstrates agonist-independent activity. Clozapine decreased the basal activity of the S267K receptor to vector control levels. Therefore, we can conclude that the S267K mutation renders the 5-HT(6) receptor constitutively active and that clozapine is an inverse agonist at the mutant 5-HT(6) receptor. These results indicate that the c-terminal region of il3 of the G(s)-coupled 5-HT(6) receptor is a key domain for G-protein coupling, similar to the G(q)-coupled 5-HT receptors. The inverse agonist action of clozapine indicates that drugs displaying competitive antagonist activity at native 5-HT(6) receptors may display inverse agonist activity at the constitutively activated form of the receptor.

Constitutive activity of G-protein coupled receptors: emphasis on serotonin receptors.

Several lines of evidence indicate that G-protein coupled receptors (GPCR) may exist in a state that allows a tonic level of stimulation in vivo (constitutive activity). Several native forms of GPCR, when expressed in recombinant cell lines, display significant signal transduction stimulation in the absence of activating ligand. Many GPCR, including three serotonin receptors, display robust constitutive activation upon the mutation of a single amino acid, indicating mutations producing inappropriate constitutive activation may be etiological factors in diseases. If constitutive activity of GPCR is as common a phenomenon as some researchers suspect, this would suggest significant alterations in the classical model of ligand-receptor interactions. One of the most significant implications of constitutive activity for pharmacologists and medicinal chemists, is the possibility of developing drugs that lower the level of constitutive activity. Such compounds have been termed inverse agonists . These drugs, in theory, would have different physiological effects, and therefore possibly different therapeutic potential, than classical competitive receptor antagonists ( neutral antagonists ). Theoretical issues concerning constitutive activity in the GPCR family and some of the evidence supporting the existence of constitutive activity in the GPCR family is reviewed. Studies are presented demonstrating the procedures for producing and characterizing constitutive activated forms of serotonin receptors, including the demonstration of inverse agonist activity of drugs on these receptors.