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Competing Interests: POTENTIAL CONFLICT OF INTEREST: The authors have no conflicts of interest to disclose.

Newborn screening (NBS) identifies infants at risk for congenital disorders for which early intervention has been shown to improve outcomes (1). State public health programs are encouraged to screen for disorders on the national Recommended Uniform Screening Panel (RUSP), which increased from 29 disorders in 2005 to 35 in 2018.* The RUSP includes hearing loss (HL) and critical congenital heart defects, which can be detected through point-of-care screening, and 33 disorders detected through laboratory screening of dried blood spot (DBS) specimens. Numbers of cases for 33 disorders on the RUSP (32 DBS disorders and HL) reported by 50 U.S. state programs were tabulated. The three subtypes of sickle cell disease (SCD) listed as separate disorders on the RUSP (S,S disease; S,beta-thalassemia; and S,C disease) were combined for the current analysis, and the frequencies of the resulting disorders were calculated relative to annual births. During 2015-2017, the overall prevalence was 34.0 per 10,000 live births. Applying that frequency to 3,791,712 live births in 2018, † approximately 12,900 infants are expected to be identified each year with one of the disorders included in the study. The most prevalent disorder is HL (16.5 per 10,000), and the most prevalent DBS disorders are primary congenital hypothyroidism (CH) (6.0 per 10,000), SCD (4.9 per 10,000), and cystic fibrosis (CF) (1.8 per 10,000). Notable changes in prevalence for each of these disorders have occurred since the previous estimates based on 2006 births (2). The number of infants identified at a national level highlights the effect that NBS programs are having on infant health through early detection, intervention, and potential improved health, regardless of geographic, racial/ethnic, or socioeconomic differences., Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. Sari Edelman, Cynthia F. Hinton, Yvonne Kellar-Guenther, Sarah McKasson, Joshua I. Miller, Jelili Ojodu, Marci K. Sontag, Sikha Singh, and Careema Yusuf report grants from Health Resources and Services Administration during the conduct of the study. No other potential conflicts of interest were disclosed.

Objectives: To measure the rates of thyroid gland imaging and levothyroxine (L-T 4 ) discontinuation and to assess whether discontinuation was monitored with thyroid-stimulating hormone testing in subjects with congenital hypothyroidism., Study Design: This is a retrospective analysis of claims data from the IBM MarketScan Databases for children born between 2010 and 2016 and continuously enrolled in a noncapitated employer-sponsored private health insurance plan or in Medicaid for ≥36 months from the date of the first filled L-T 4 prescription., Results: There were 263 privately insured and 241 Medicaid-enrolled children who met the inclusion criteria. More privately insured than Medicaid-enrolled children had imaging between the first filled prescription and 180 days after the last filled prescription (24.3% vs 12.9%; P = .001). By 36 months, 35.7% discontinued L-T 4 , with no difference by insurance status (P = .48). Among those who discontinued, 29.1% of privately insured children and 47.7% of Medicaid-enrolled children had no claims for thyroid-stimulating hormone testing within the next 180 days (P = .01)., Conclusions: Nearly one-third of children with suspected congenital hypothyroidism discontinued L-T 4 by 3 years and fewer Medicaid-enrolled than privately insured children received timely follow-up thyroid-stimulating hormone testing. Future studies are indicated to understand the quality of care and developmental outcomes for children with congenital hypothyroidism and barriers to guideline adherence in evaluating for transient congenital hypothyroidism., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Preparedness planning is essential to minimizing the impact of disasters on communities and individuals. Attention to the needs of people with disabilities is vital as they have additional needs before, during and after a disaster that are specific to the disabling condition. In this Commentary, we emphasize national guidelines on disability inclusion in emergency preparedness. We examine some potential areas of planning and response that need attention as suggested by preparedness data for people with self-reported disabilities (also referred to as access and functional needs) and highlight selected resources (e.g., tools, trainings, and online webinars) to enhance whole community preparedness and disability inclusion efforts. This Commentary intends to bridge the gap between those various facets of preparedness, at all levels of government and among individuals, with the aim of ensuring that the whole community is prepared to adapt, withstand and rapidly recover from disruptions due to disasters., (Published by Elsevier Inc.)

Newborn screening (NBS) identifies infants with rare conditions to prevent death or the onset of irreversible morbidities. Conditions on the Health and Human Services Secretary's Recommended Uniform Screening Panel have been adopted by most state NBS programs, providing a consistent approach for identification of affected newborns across the United States. Screen-positive newborns are identified and referred for confirmatory diagnosis and follow-up. The designation of a clinically significant phenotype precursor to a clinical diagnosis may vary between clinical specialists, resulting in diagnostic variation. Determination of disease burden and birth prevalence of the screened conditions by public health tracking is made challenging by these variations. This report describes the development of a core group of new case definitions, along with implications, plans for their use, and links to the definitions that were developed by panels of clinical experts. These definitions have been developed through an iterative process and are piloted in NBS programs. Consensus public health surveillance case definitions for newborn screened disorders will allow for consistent categorization and tracking of short- and long-term follow-up of identified newborns at the local, regional, and national levels., Competing Interests: Conflicts of Interest: The authors declare no conflict of interest.

Introduction: Approximately 40 million people in the U.S. identify as having a serious disability, and people with disabilities experience many health disparities compared with the general population. The Guide to Community Preventive Services (The Community Guide) identifies evidence-based programs and policies recommended by the Community Preventive Services Task Force (Task Force) to promote health and prevent disease. The Community Guide was assessed to answer the questions: are Community Guide public health intervention recommendations applicable to people with disabilities, and are adaptations required?, Methods: An assessment of 91 recommendations from The Community Guide was conducted for 15 health topics by qualitative analysis involving three data approaches: an integrative literature review (years 1980-2011), key informant interviews, and focus group discussion during 2011., Results: Twenty-six recommended interventions would not need any adaptation to be of benefit to people with disabilities. Forty-one recommended interventions could benefit from adaptations in communication and technology; 33 could benefit from training adaptations; 31 from physical accessibility adaptations; and 16 could benefit from other adaptations, such as written policy changes and creation of peer support networks. Thirty-eight recommended interventions could benefit from one or more adaptations to enhance disability inclusion., Conclusions: As public health and healthcare systems implement Task Force recommendations, identifying and addressing barriers to full participation for people with disabilities is important so that interventions reach the entire population. With appropriate adaptations, implementation of recommendations from The Community Guide could be successfully expanded to address the needs of people with disabilities., (Published by Elsevier Inc.)

Background: We assessed sociodemographic and clinical factors that are associated with survival among infants with congenital diaphragmatic hernia (CDH)., Methods: Using data from the Metropolitan Atlanta Congenital Defects Program, we ascertained 150 infants born with CDH between 1979 and 2003 and followed via linkage with state vital records and the National Death Index. Kaplan-Meier survival probabilities and adjusted hazard ratios (HRs) were calculated for socioeconomic and clinical characteristics., Results: Survival increased from 40 to 62% over the study period. White infants born before 1988 were 2.9 times less likely to survive than those born after 1988. Black infants' survival did not show significant improvement after 1988. White infants' survival was not significantly affected by poverty, whereas black infants born in higher levels of poverty were 2.7 times less likely to survive than black infants born in lower levels of neighborhood poverty. White infants with multiple major birth defects were 2.6 times less likely to survive than those with CDH alone. The presence of multiple defects was not significantly associated with survival among black infants., Conclusions: Survival among infants and children with CDH has improved over time among whites, but not among blacks. Poverty is associated with lower survival among blacks, but not among whites. The presence of multiple defects is associated with lower survival among whites, but not among blacks. The differential effects of poverty and race should be taken into account when studying disparities in health outcomes. Birth Defects Research 109:816-823, 2017. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Unlabelled: Newborn screening (NBS) is intended to identify congenital conditions prior to the onset of symptoms in order to provide early intervention that leads to improved outcomes. NBS is a public health success, providing reduction in mortality and improved developmental outcomes for screened conditions. However, it is less clear to what extent newborn screening achieves the long-term goals relating to improved health, growth, development and function. We propose a framework for assessing outcomes for the health and well-being of children identified through NBS programs. The framework proposed here, and this manuscript, were approved for publication by the Secretary of Health and Human Services' Advisory Committee on Heritable Disorders in Newborns and Children (ACHDNC). This framework can be applied to each screened condition within the Recommended Uniform Screening Panel (RUSP), recognizing that the data elements and measures will vary by condition. As an example, we applied the framework to sickle cell disease and phenylketonuria (PKU), two diverse conditions with different outcome measures and potential sources of data. Widespread and consistent application of this framework across state NBS and child health systems is envisioned as useful to standardize approaches to assessment of outcomes and for continuous improvement of the NBS and child health systems., Significance: Successful interventions for newborn screening conditions have been a driving force for public health newborn screening for over fifty years. Organizing interventions and outcome measures into a standard framework to systematically assess outcomes has not yet come into practice. This paper presents a customizable outcomes framework for organizing measures for newborn screening condition-specific health outcomes, and an approach to identifying sources and challenges to populating those measures., (Published by Elsevier Inc.)

Routine second screening of most newborns at 8-14 days of life for a panel of newborn conditions occurs in 12 U.S. states, while newborns in the other states typically undergo only a single routine newborn screen. The study objective was to evaluate screening consequences for primary congenital hypothyroidism (CH) in one- and two-screen states according to laboratory practices and medical or biochemical characteristics of screen-positive cases. Individual-level medical and biochemical data were retrospectively collected and analyzed for 2251 primary CH cases in one-screen (CA, WI) and two-screen (AL, DE, MD, OR, TX) states. Aggregate data were collected and analyzed for medical and biochemical characteristics of all screened newborns in the states. Among the states evaluated in this study, the detection rate of primary CH was higher in the one-screen states. In the two-screen states, 11.5% of cases were detected on the second screen. In multivariate analyses, only race/ethnicity was a significant predictor of cases identified on the first versus second screen, which likely reflects a physiologic difference in primary CH presentation. Newborn screening programs must heed the potential for newborns with CH not being detected by a single screen, particularly newborns of certain races/ethnicities. If the two-screen states converted to a single screen using their current algorithms, newborns currently identified on the routine second screen would presumably not be detected, resulting in probable delayed diagnosis and treatment. However, based on the one-screen state experiences, with appropriate modifications in screening method and algorithm, the two-screen states might convert to single screen operation for CH without loss in performance., (Published by Elsevier Inc.)

There is no clear consensus among state newborn screening programs on whether routine second screening of newborns identifies clinically relevant cases of congenital adrenal hyperplasia. This retrospective study evaluated laboratory practices, along with biochemical and medical characteristics of congenital adrenal hyperplasia (CAH) cases (1) detected on the first newborn screen in one-screen compared to two-screen states, and (2) detected on the first versus the second screen in the two-screen states, to determine the effectiveness of a second screen. A total of 374 confirmed cases of CAH from 2 one-screen states and 5 two-screen states were included in this study. Demographic data and diagnostic information on each reported case were collected and analyzed. Additionally, laboratory data, including screening methodologies and algorithms, were evaluated. The one-screen states reported 99 cases of CAH out of 1,740,586 (1 in 17,500) newborns screened: 88 (89%) identified on the first screen and 5 (5%) identified on the targeted second screen. The two-screen states reported 275 cases of CAH out of 2,629,627 (1 in 9500) newborns screened: 165 (60%) identified on the first screen and 99 (36%) identified on the second screen. Using a multivariate model, the only significant predictor of whether a case was identified on the first or the second screen in the two-screen states was the type of CAH. Compared with classical salt-wasting CAH, classical simple virilizing and non-classical CAH cases were less likely to be detected on the first versus the second screen. The routine second newborn screen is important for identifying children with CAH, particularly simple virilizing and non-classical forms, which might otherwise not be captured through a single screen., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Recent public health emergencies including Hurricane Katrina (2005), the influenza H1N1 pandemic (2009), and the Ebola virus disease outbreak in West Africa (2014–2015) have demonstrated the importance of multiple-level emergency planning and response. An effective response requires integrating coordinated contributions from community-based health care providers, regional health care coalitions, state and local health departments, and federal agency initiatives. This is especially important when planning for the needs of children, who make up 23% of the U.S. population (1) and have unique needs that require unique planning strategies.

Purpose: The aim of this study was to describe the methods, cases, and initial results of a pilot project using existing public health data collection programs (birth defect surveillance or newborn screening) to conduct long-term follow-up of children with metabolic disorders., Methods: California, Iowa, New York, and Utah expanded birth defect surveillance or newborn screening programs to collect long-term follow-up data on 19 metabolic disorders. Data elements to monitor health status and services delivered were identified, and record abstraction and data linkages were conducted. Children were followed up through to the age of 3 years., Results: A total of 261 metabolic cases were diagnosed in 1,343,696 live births (19.4 cases/100,000; 95% confidence interval = 17.1-21.8). Four deaths were identified. Children with fatty acid oxidation disorders had a higher percentage of health service encounters compared with children with other disorders of at least one health service encounter (hospitalization, emergency room, metabolic clinic, genetic service provider, or social worker) except for hospitalizations; children with organic acid disorders had a higher percentage of at least one hospitalization during their third year of life than children with other disorders., Conclusion: Existing public health data programs can be leveraged to conduct population-based newborn screening long-term follow-up. This approach is flexible according to state needs and resources. These data will enable the states in assessing health burden, assuring access to services, and supporting policy development.

Objective: Critical congenital heart disease (CCHD) was recently added to the U.S. Recommended Uniform Screening Panel for newborns. This evaluation aimed to estimate screening time and hospital cost per newborn screened for CCHD using pulse oximetry as part of a public health economic assessment of CCHD screening., Methods: A cost survey and time and motion study were conducted in well-newborn and special/intensive care nurseries in a random sample of seven birthing hospitals in New Jersey, where the state legislature mandated CCHD screening in 2011. The sample was stratified by hospital facility level, hospital birth census, and geographic location. At the time of the evaluation, all hospitals had conducted CCHD screening for at least four months., Results: Mean screening time per newborn was 9.1 (standard deviation = 3.4) minutes. Hospitals' total mean estimated cost per newborn screened was $14.19 (in 2011 U.S. dollars), consisting of $7.36 in labor costs and $6.83 in equipment and supply costs., Conclusions: This federal agency-state health department collaborative assessment is the first state-level analysis of time and hospital costs for CCHD screening using pulse oximetry conducted in the U.S. Hospitals' cost per newborn screened for CCHD with pulse oximetry is comparable with cost estimates of existing newborn screening tests. Hospitals' equipment costs varied substantially based on the pulse oximetry technology employed, with lower costs among hospitals that used reusable screening sensors. In combination with estimates of screening accuracy, effectiveness, and avoided costs, information from this evaluation suggests that CCHD screening is cost-effective.

Background and Objective: New Jersey was the first state to implement legislatively mandated newborn pulse oximetry screening (POxS) in all licensed birthing facilities to detect critical congenital heart defects (CCHDs). The objective of this report was to evaluate implementation of New Jersey's statewide POxS mandate., Methods: A 2-pronged approach was used to collect data on infants screened in all New Jersey birthing facilities from August 31, 2011, through May 31, 2012. Aggregate screening results were submitted by each birthing facility. Data on failed screens and clinical characteristics of those newborns were reported to the New Jersey Birth Defects Registry (NJBDR). Three indicators were used to distinguish the added value of mandated POxS from standard clinical care: prenatal congenital heart defect diagnosis, cardiology consultation or echocardiogram indicated or performed before PoxS, or clinical findings at the time of POxS warranting a pulse oximetry measurement., Results: Of 75,324 live births in licensed New Jersey birthing facilities, 73,320 were eligible for screening, of which 99% were screened. Forty-nine infants with failed POxS were reported to the NJBDR, 30 of whom had diagnostic evaluations solely attributable to the mandated screening. Three of the 30 infants had previously unsuspected CCHDs and 17 had other diagnoses or non-CCHD echocardiogram findings., Conclusions: In the first 9 months after implementation, New Jersey achieved a high statewide screening rate and established surveillance mechanisms to evaluate the unique contribution of POxS. The screening mandate identified 3 infants with previously unsuspected CCHDs that otherwise might have resulted in significant morbidity and mortality and also identified other significant secondary targets such as sepsis and pneumonia.

Objective: To implement a 6-month quality improvement project in 15 primary care pediatric practices to improve short-term newborn screening (NBS) follow-up., Methods: At the start of the project, each practice completed a survey to evaluate office systems related to NBS and completed a chart audit. Practice teams were provided information about NBS and trained in quality-improvement methods, and then implemented changes to improve care. Monthly chart audits over a 6-month period were completed to assess change., Results: At baseline, almost half of practices completed assessment of infants for NBS; after 6 months, 80% of practices completed assessment of all infants. Only 2 practices documented all in-range results and shared them with parents at baseline; by completion, 10 of 15 practices documented and shared in-range results for ≥ 70% of infants. Use of the American College of Medical Genetics ACTion sheets, a decision support tool, increased from 1 of 15 practices at baseline to 7 of 15 at completion., Conclusions: Practices were successful in improving NBS processes, including assessment, documentation, and communication with families. Providers perceived no increase in provider time at first visit, 2- to 4-week visit, or during first contact with the family of an infant with an out-of-range result after implementation of improved processes. Primary care practices increased their use of decision support tools after the project.

The US Secretary of Health and Human Services' Advisory Committee on Heritable Disorders in Newborns and Children provides guidance on reducing the morbidity and mortality associated with heritable disorders detectable through newborn screening. Efforts to systematically evaluate health outcomes, beyond long-term survival, with a few exceptions, are just beginning. To facilitate these nascent efforts, the US Secretary of Health and Human Services' Advisory Committee on Heritable Disorders in Newborns and Children initiated a project to define the major overarching questions to be answered to assure that newborn screening is meeting its goal of achieving the best quality outcome for the affected children and their families. The questions identified follow the central components of long-term follow-up-care coordination, evidence-based treatment, continuous quality improvement, and new knowledge discovery-and are framed from the perspectives of the state and nation, primary and specialty healthcare providers, and the impacted families. These overarching questions should be used to guide the development of long-term follow-up data systems, quality health indicators, and specific data elements for evaluating the newborn screening system.

The prevalence of hemoglobinopathies differs among populations due to genetic differences and due to the protective effects of the heterozygote (carrier) state against malaria. Because of the difference in genetic distribution, public health programs have weighed the ethical versus practical implications of ethnically targeted versus universal newborn, and where applicable, prenatal screening. We examine newborn and prenatal screening for hemoglobinopathies in relation to the use of 'race' and ethnicity to assess risk for genetic conditions. First, categories of race/ethnicity are social constructs, therefore, observed or self-identified broad racial/ethnic categories are correlated but not necessarily reliable indicators of geographic ancestry or genetic risk. Second, targeting based on ethnicity poses serious issues of logistics and equity for public health programs and clinical services. In the past, newborn screening for hemoglobinopathies in the United States and United Kingdom was often selective, targeted to women of certain ethnic groups or areas with large concentrations of ethnic minority groups. Presently, newborn screening for hemoglobinopathies is universal in both countries and programs emphasize that individuals of all ethnic backgrounds are at risk for carrying a hemoglobin genetic variant. Reported race/ethnicity is still used as a criterion for offering prenatal carrier testing in the United States, where it is not a public health responsibility. In the United Kingdom, prenatal screening under the National Health Service is universal in high-prevalence areas and in low-prevalence areas is targeted based on reported ancestry. The continued use of targeted prenatal screening in both countries reflects the different purposes and modes of laboratory testing in newborn and prenatal screening. The ethical imperative to identify as many affected infants with life-threatening conditions as possible in newborn screening programs is not applicable to prenatal carrier testing. Because newborn screening dried blood spot specimens are tested for multiple disorders, targeted screening poses serious logistical challenges which is not the case in prenatal screening.

The reported incidence rate of primary congenital hypothyroidism (CH) has been increasing in the United States over the past 2 decades. We have considered the possibility that the inclusion of cases of transient hypothyroidism has inflated the reported incidence rate of CH. Assessing the effects of cases of transient hypothyroidism on the incidence rate is problematic, because the definitions, diagnostic criteria, and differentiation from transient hyperthyrotropinemia vary widely among state newborn screening programs. Among the 4 etiologies for transient hypothyroidism (maternal thyrotropin receptor-blocking antibodies, exposure to maternal antithyroid medications, iodine deficiency, and iodine excess), there is little evidence of increases in the incidence rate from thyrotropin receptor-blocking antibodies. Exposure to antithyroid drugs could contribute significantly to the incidence rate of transient CH, given the high estimated incidence of active maternal hyperthyroidism. Iodine deficiency or excess in the United States seems unlikely to have contributed significantly to the incidence rate of CH, because the secular trend toward lower iodine intake among women of reproductive age in the 1980s and 1990s seems to have plateaued, and perinatal iodine exposure has presumably declined as a result of recommendations to discontinue using iodine-containing disinfectants. Although the female-to-male sex ratio among newborns with thyroid agenesis or dysgenesis (the most common causes of CH) is typically 2:1, analysis of the sex ratio of newborns diagnosed with presumed CH in the United States suggests that a substantial proportion might have transient hypothyroidism or hyperthyrotropinemia, because the sex ratio has been well below the expected 2:1 ratio. Combined ultrasonography and (123)I scintigraphy of the thyroid gland are effective tools for identifying cases of thyroid agenesis and dysgenesis and can help to differentiate cases of transient hypothyroidism from true CH. Imaging is also a vital component in evaluating children who, at 3 years of age, undergo a trial of discontinuation of levothyroxine treatment to test for persistence of hypothyroidism. Ultimately, thyroid gland imaging, in conjunction with long-term follow-up studies that appropriately assess and report whether there was permanence of hypothyroidism, will be necessary to address the true incidence rate of CH and any contribution to the observed rate by transient cases of hypothyroidism or hyperthyrotropinemia.

Primary congenital hypothyroidism (CH) is a common and preventable cause of intellectual disability. The incidence rate of CH has been reported to be increasing in the United States, but the factors behind the observed rate increase are not known. We summarize here the data presented at a workshop on CH, at which factors potentially related to the CH-incidence-rate increase (namely, race, ethnicity, sex, and birth outcomes) were evaluated. Data sources for the analyses included a national data set of newborn-screening results and state-specific data from newborn-screening programs in California, Massachusetts, New York, and Texas. The incidence rate of CH increased in the United States by 3% per year; however, an increase did not occur in all states, at a constant rate, or even at the same rate. Analysis of US data (1991-2000) showed a CH-incidence-rate increase only among white newborns. More recently, in California (2000-2007), the rate was constant in non-Hispanic newborns, but it increased among Hispanic newborns. In the national data, the CH-incidence rate increased similarly among boys and girls, whereas in Texas (1992-2006), the rate among boys increased significantly more than among girls and varied according to race and ethnicity. In Massachusetts (1995-2007), low birth weight newborns or newborns who had a delayed rise in thyrotropin concentration accounted for the majority of the recent rate increase. Race, ethnicity, sex, and pregnancy outcomes have affected the observed increasing incidence rate of CH, although there have been some inconsistencies and regional differences. The association with preterm birth or low birth weight could reflect the misclassification of some cases of transient hypothyroxinemia as true CH. Future studies of risk factors should focus on correct initial identification and reporting of demographic characteristics and pregnancy outcomes for cases of CH. In addition, long-term follow-up data of presumed cases of CH should be ascertained to differentiate true cases of CH from cases of transient hypothyroidism.

Patch-clamp methodology was applied to principal cell apical membranes of rabbit cortical collecting tubule (CCT) primary cultures grown on collagen supports in the presence of aldosterone (1.5 microM). The most frequently observed channel had a unit conductance of 3-5 pS, nonlinear current-voltage (I-V) relationship, Na permeability (PNa)-to-K permeability (PK) ratio greater than 19:1, and inward current at all applied potentials (Vapp) less than +80 mV (n = 41). Less frequently, an 8- to 10-pS channel with a linear I-V curve, PNa/PK less than 5:1, and inward current at Vapp less than +40 mV was also observed (n = 7). Luminal amiloride (0.75 microM) decreased the open probability (Po) for both of these channels. Mean open time for the high-selectivity Na+ channel was 2.1 +/- 0.5 s and for the low-selectivity Na+ channel was 50 +/- 12 ms. In primary cultures grown without aldosterone the high-selectivity Na+ channel was rarely observed (1 of 32 patches). Lastly, a 26- to 35-pS channel, nonselective for Na+ over K+, was not activated by cytoplasmic Ca2+ or voltage nor inhibited by amiloride (n = 17). We conclude that under specific growth conditions, namely permeable transporting supports and chronic mineralocorticoid hormone exposure, principal cell apical membranes of rabbit CCT primary cultures contain 1) both high-selectivity and low-selectivity, amiloride-inhibitable Na+ channels and 2) amiloride-insensitive, nonselective cation channels.

Rabbit cortical collecting tubule (RCCT) primary cultures, were grown on permeable, collagen supports with 1.5 microM aldosterone. Single K+ permeable channels in principal cell apical membranes were examined. At applied patch pipette potential (Vapp) from -60 to +60 mV (cell interior with respect to pipette interior), outward currents (cell to pipette) with a unitary conductance of 8 to 10 pS were seen in cell-attached (N = 31) and excised inside-out (N = 15) patches. At resting membrane potential (Vapp = 0 mV), mean open probability (Po = 0.85 +/- 0.16) decreased by 50% with 0.75 mM luminal BaCl2 exposure. In cell-attached patches, a second type of outward current was seen only at extreme depolarization, Vapp greater than +80 mV (N = 9). Usually in the closed state (Po less than 0.0005) at no applied potential, Po for this 150 pS channel increased dramatically with depolarization and/or raising cytoplasmic Ca2+. With a calculated K+ equilibrium potential of -84 mV, excised patch reversal potentials were less than -50 mV for both the above channel types, indicating high selectivity for K+ over Na+. In cultures grown without aldosterone low conductance K+ channels were rarely observed, while mineralocorticoid status did not appear to affect high conductance K+ channel frequency. Finally, a 30 pS cation channel was found to be nonselective for K+ over Na+, and insensitive to voltage, intracellular Ca2+ or luminal Ba2+. We conclude that: 1) Principal cell apical membranes from aldosterone-stimulated, RCCT primary cultures contain (a) low conductance, Ba(2+)-inhibitable and (b) high conductance, Ca2+/voltage-dependent K+ channels; and c) nonselective cation channels. 2) The low conductance K+ channel may play an important physiologic role in native RCCT mineralocorticoid-controlled K+ secretion, while the latter two channels' functions are unknown, although similar channels have been suggested to play a role in cell volume regulation.

The first step in net active transepithelial transport of sodium in tight epithelia is mediated by the amiloride-blockable sodium channel in the apical membrane. This sodium channel is the primary site for discretionary control of total body sodium and, therefore, investigating its regulatory mechanisms is important to our understanding of the physiology of fluid and electrolyte balance. Because essentially all of the regulatory sites on the channel are on the intracellular surface, patch clamp methods have proven extremely useful in the electrophysiological characterization of the sodium channel by isolating it from other channel proteins in the epithelial membrane and by allowing access to the intracellular surface of the protein. We have examined three different regulatory mechanisms. (1) Inhibition of channel activity by activation of protein kinase C; (2) activation of the channel by agents which activate G-proteins; and (3) modulation of channel kinetics and channel number by mineralocorticoids. Activation of protein kinase C by phorbol esters or synthetic diacylglycerols reduces the open probability of sodium channels. Protein kinase C can be activated in a physiological context by enhancing apical sodium entry. Actions which reduce sodium entry (low luminal sodium concentrations or the apical application of amiloride) increase channel open probability. The link between sodium entry and activation of protein kinase C appears to be mediated by intracellular calcium activity linked to sodium via a sodium/calcium exchange system. Thus, the intracellular sodium concentration is coupled to sodium entry in a negative feedback loop which promotes constant total entry of sodium. Activation of G-proteins by pertussis toxin greatly increases the open probability of sodium channels. Since channels can also be activated by pertussis toxin or GTP gamma S in excised patches, the G-protein appears to be closely linked in the apical membrane to the sodium channel protein itself. The mechanism for activation of this apical G-protein, when most hormonal and transmitter receptors are physically located on the basolateral membrane, is unclear. Mineralocorticoids such as aldosterone have at least two distinct effects. First, as expected, increasing levels of aldosterone increase the density of functional channels detectable in the apical membrane. Second, contrary to expectations, application of aldosterone increases the open probability of sodium channels. Thus aldosterone promotes the functional appearance of new sodium channels and promotes increased sodium entry through both new and pre-existant channels.

The receptors mediating adrenergic stimulation of acid secretion by Amphiuma jejunum were characterized in this study using alpha- and beta-adrenergic agonists and antagonists. Isolated segments of jejunum were mounted in Ussing chambers and bathed in Cl- -free (SO4(2-] medium. Shortcircuit current (Isc) and acid secretion (JH) were recorded, the latter by measuring the rate of alkalinization of the serosal medium. The beta-adrenergic receptor antagonist, propranolol (10(-4) M), had no effect on the Isc and JH stimulated by norepinephrine (NE). The alpha 2-adrenergic agonists, clonidine and UK-14,304, mimicked the effect of NE, with effective concentrations providing 50% maximal delta Isc of 2.0 X 10(-7) and 9.0 X 10(-8) M, respectively. NE added subsequently produced no greater stimulation. In contrast, the alpha 1-adrenergic agonists, phenylephrine and methoxamine, produced little stimulation of JH and Isc; NE added subsequently stimulated the Isc. The alpha 1-adrenergic antagonist prazosin had no effect on the NE-induced Isc or JH, whereas the alpha 2-adrenergic antagonist yohimbine inhibited the NE-stimulated Isc with a half-maximal effective concentration of 3.5 X 10(-7) M. Yohimbine (10(-4) M) reduced the NE-stimulated Isc by 88%, whereas the spontaneous Isc was reduced by only 12%. These results demonstrate that alpha 2-adrenergic receptors on the basolateral membrane of Amphiuma enterocytes mediate NE-enhanced, but not spontaneous, intestinal acid secretion.

This report describes the expression of a sodium-selective, amiloride-blockable conductance in Xenopus oocytes that have been injected with RNA prepared from a distal nephron cell line (A6). After injecting the RNA into mature oocytes (stage V or VI) and incubating the oocytes for 2-4 days, the oocytes were examined for amiloride-blockable current. The RNA induced a substantial amiloride-blockable current. Uninjected or water-injected oocytes had no measurable amiloride-blockable current. RNA prepared from aldosterone-treated A6 cells was much more effective in inducing amiloride-blockable sodium current than RNA prepared from aldosterone-depleted A6 cells. Oocytes injected with RNA prepared from mineralocorticoid-depleted cells appeared very similar to water-injected oocytes. The amiloride-blockable current in oocytes has a reversal potential of approximately +50 - +60 mV, which varies 61 mV/decade change in external sodium concentration, suggesting that the current is highly selective for sodium over other ions. In addition, the concentration of amiloride that produces half block of the current is 48 +/- 8 nM. Thus the current expressed in oocytes appears very similar to sodium-selective currents observed from the apical membranes of various tight epithelial tissues.

Catecholamines stimulate Na+-dependent acid secretion by Amphiuma small intestine. Studies were undertaken to localize the response within the mucosa and characterize the effect on Na+ and Cl- transport. Stripped segments of jejunum were mounted in tissue chambers that permitted isolation of villus or intervillus epithelium. In Cl-free medium, norepinephrine (NE) stimulated the transepithelial voltage (Vms) in both villus and intervillus epithelium, whereas galactose and valine elevated Vms predominately in the villus. Paired segments of whole mucosa were maintained under short circuit while the rate of acid secretion (JH) was measured by titration of the unbuffered serosal medium and unidirectional fluxes of Na+ were measured by 22Na. NE significantly stimulated net Na+ absorption (JNanet), short circuit current (Isc), and JH. Amiloride reduced JH and Isc in NE-stimulated tissues and blocked the stimulation of JNanet by NE. The NE-induced current was nearly completely and reversibly inhibited by replacement of luminal medium HCO3- or CO2. NE significantly stimulated net Cl- absorption without changing Isc or JH. It is concluded that cells throughout the mucosa respond to catecholamines with enhanced Na+ and Cl- absorption, possibly through induction or stimulation of Na+-H+ and Cl- -HCO3- exchange.