Glutamine transport in human and rat placenta.

Glutamine plays an important role in fetal nutrition. This study explored the transport of [3H]glutamine into apical and basal predominant membrane vesicles derived from rat and human placenta. Na+-dependent glutamine transport was present in both apical and basal predominant vesicles derived from 20- and, to a lesser degree, 14-day gestation rat placenta. Amino-acid transport systems A, ASC-like, B(o,+) (in apical membrane vesicles) and, perhaps, y+L were involved in Na+-dependent glutamine transport. Na+-dependent glutamine uptake into human placental microvillus and basolateral membrane vesicles also occurred via several distinct transport activities. Glutamine transport via system N was not detected in either rat or human placental preparations. Na+-dependent glutamine transport in the rat was more pronounced in basal as compared to apical membrane vesicles. Conversely, in the human preparations, activity was significantly higher in microvillus as compared to basolateral membrane vesicles. It is concluded that Na+-dependent glutamine transport occurs through a variety of transport agencies in both the rat and human placenta. Transport varies with ontogeny and between species.

Anionic amino acid transporter expression in late gestation rodent yolk sac.

The yolk sac plays an important role in fetal nutrition. Transport of amino acids by the rodent visceral yolk sac has been shown previously. We have demonstrated the presence of several amino acid transport proteins capable of the Na(+)-dependent transport of anionic amino acids within late gestation mouse visceral yolk sac and uterine epithelium. We speculate that these proteins may be involved in the efflux of glutamate from the fetal to the maternal circulations.

Ontogeny of amino acid transport system A in rat placenta.

Amino acid transport System A has previously been demonstrated in apical membranes derived from rat placenta, as well as in apical and basal membranes derived from human placenta. We have studied Na(+)-dependent alpha-(methylamino)isobutyric acid (MeAIB) transport in apical and basal predominant membrane fractions prepared from 14 and 20 day gestation rat placenta. Marker enzyme recoveries did not differ significantly between age groups. Markers for intracellular organelles were also found to be comparable. Na(+)-dependent MeAIB transport was not sensitive to freezing and could be found in all membrane components tested. Kinetic parameters were studied--Km = 852 +/- 215 microM, Vmax = 718 +/- 126 pmol/5 sec/mg protein--20 day apical; Km = 748 +/- 269 microM, Vmax = 610 +/- 176 pmol/5 sec/mg protein--20 day basal-predominant; Km 614 +/- 261 microM, Vmax = 123 +/- 45 pmol/5 sec/mg protein-14 day apical. Kinetic parameters could not be determined in the 14 day gestation basal-predominant fraction because of the small amount of uptake present. We conclude that System A like activity is found in both apical and basal predominant membrane fractions derived from rat placenta, and that this activity increases over the last one third of gestation.

Placental anionic and cationic amino acid transporter expression in growth hormone overexpressing and null IGF-II or null IGF-I receptor mice.

The role of growth hormone (GH), insulin-like growth factor (IGF)-II and the IGF-I receptor (IGF-Ir) in the regulation of the in vivo expression of Na(+)-coupled anionic [System X-AG; GLAST1 (EAAT1), GLT1 (EAAT2), EAAC1 (EAAT3), EAAT4; where the human homologues of amino acid transport proteins first cloned in the rat are given in parentheses] and Na(+)-independent cationic (System y(+);CAT1) amino acid transport proteins was evaluated by comparing transporter expression in day 17 placentae of mice that overexpressed bovine GH (GH+) or that carried null gene mutations for IGF-II or IGF-Ir. Northern analysis revealed no apparent difference in the mRNA content of GLAST1 (EAAT1), EAAC1 (EAAT3), or EAAT4, in homogenates of GH+ placentae, but levels of GLT1 (EAAT2) and CAT1 mRNA were increased. Immunoblot analysis revealed that whole-placental steady-state GLAST1 (EAAT1), EAAC1 (EAAT3), and EAAT4 protein levels were not affected by GH+, whereas GLT1 (EAAT2) levels were increased. Immunohistochemical analysis showed that the cell-specific expression of the anionic and CAT1 transporters was not affected by overexpression of GH. Similar analyses of null IGF-II placentae demonstrated increases in GLAST1 (EAAT1), EAAT4 and CAT1 mRNAs. Parallel immunoblot analysis demonstrated decreased expression of GLT1 (EAAT2), GLAST1 (EAAT1) and EAAC1 (EAAT3) protein, but an increased expression of EAAT4. In null IGF-II and IGF-Ir placentae, however, GLT1 (EAAT2) and EAAC1 (EAAT3) protein content was decreased in junctional zone cells, whereas CAT1 content was increased in junctional and labyrinth zone cells. These data indicate that an excess level of GH stimulates GLT1 (EAAT2) expression and that a normal level of IGF-II is required for typical expression of GLT1 (EAAT2), GLAST1 (EAAT1) and EAAC1 (EAAT3), but that IGF-II downregulates the expression of EAAT4 and CAT1.

Demonstration of system y+L activity on the basal plasma membrane surface of rat placenta and developmentally regulated expression of 4F2HC mRNA.

Na(+)-independent cationic amino acid transport in the rat placenta occurs by leucine-sensitive and leucine-insensitive pathways. The ontogeny of these transport mechanisms within the rat placenta has been described recently. To assign the leucine-inhibitable portion of uptake definitively the uptake of [3H]arginine was studied in the presence of both BCH (to inhibit system Bo,+) and varied concentrations of leucine. Uptake of arginine into basal-enriched membrane vesicles derived from rat placenta was, in the presence of sodium, inhibited by micromolar concentrations of leucine, consistent with assignment of this activity to system y+L. In contrast, the majority of arginine uptake into apical-enriched membrane vesicles was leucine insensitive. Messenger RNA derived from rat placenta at days 14, 16, 18 and 20 of gestation was hybridized with full-length rat cDNA probes against NBAT and 4F2HC (thought to encode proteins associated with system bo,+ and y+L activities, respectively). No NBAT mRNA was detected, whereas 4F2HC mRNA was present at all gestational stages, increasing 12-fold over the last third of gestation. It is concluded that system y+L is present in the basal plasma membrane of the rat placenta syncytium and is subject to developmental regulation by a mechanism that alters the steady content of 4F2HC mRNA.

Effect of chronic cocaine administration on amino acid uptake in rat placental membrane vesicles.

This study evaluated the effects of chronic exposure to cocaine during pregnancy on amino acid uptake in placental membrane vesicles. Pregnant rats received 62 mg/kg of cocaine hydrochloride by intraperitoneal (IP) injection as a divided daily dose on gestation days 8-19 inclusive. Fetal body weights were significantly decreased by 19% in the cocaine group, while placental weights were unchanged. Placental apical membrane vesicles were prepared from control and cocaine-treated animals, and marker enzyme enrichments for alkaline phosphatase and [3H]-dihydroalprenolol binding did not differ between cocaine and control groups. Rates of uptake (10 sec) of selected radiolabeled amino acids were measured utilizing a rapid filtration technique. Na(+)-dependent apical membrane [3H]-glutamine transport (50 microM) was reduced by 95% (p < 0.05) in cocaine-treated compared to control placentas. Uptake of 50 microM [3H]-methyl aminoisobutyric acid (MeAIB) into apical membranes was also decreased by 43% (p < 0.05) in cocaine membranes. Na(+)-independent [3H]-arginine transport (10 microM), however, did not differ between control or cocaine-treated groups. In summary, chronic cocaine administration selectively inhibited the transport of glutamine and MeAIB into apical membrane vesicles, but had minimal effect on arginine transport. We postulate that this diminution in uptake may contribute to the fetal growth retardation noted in our model.

Dietary nucleotides affect hepatic growth and composition in the weanling mouse.

The effect of dietary nucleotides upon hepatic growth and composition was examined in weanling mice. For 5 weeks, mice were fed either Purina Rat Chow, a nucleotide-free diet (NT-), a nucleotide-free diet supplemented with a mixture of five nucleotides (0.21% w/w), (NT+) or a nucleotide-free diet supplemented with adenosine 5'-monophosphate (0.0425% w/w) (NTA). Hepatic cholesterol and lipid phosphorous were significantly higher, whereas liver weight (expressed as a percentage of body weight), and glycogen were lower in animals fed NT- vs all other groups. NTA-fed animals presented a greater contrast to the NT- group than did animals fed the mixture of nucleotides. Liver fatty acid composition and distribution of phospholipid subclasses were not affected by dietary nucleotide supplementation. Dietary nucleotide supplementation in weanling mice affects hepatic growth and composition; adenosine 5'-monophosphate may play a unique role in these effects.

Gastroesophageal reflux in the preterm infant.

Gastroesophageal reflux is a problem familiar to most pediatricians. The focus of this article is to provide a synopsis of the current state of knowledge regarding gastroesophageal reflux in the premature population, as well as to provide the practitioner with a rational basis upon which to diagnose and treat gastroesophageal reflux in this population.

Older adults' descriptions of their role expectations of nursing.

A qualitative descriptive approach was used to identify 28 older adults' role expectations of nursing. This study describes elders' opinions, expectation and health care preferences which influence their judgments about nursing. Three categories emerged from analyzing transcripts of the audiotaped interviews. The older adults expected the nurse to be knowledgeable, caring and attentive by: demonstrating professional competence when recognizing patients' needs, as well as being concerned for the individual in the responsive delivery of services. The results of this study are congruent with descriptions of valued nursing behaviors that have been identified in previous studies. Additional findings, also corroborated by prior research, indicated that nursing roles and actions may not be accurately perceived or understood by the older consumer. Nurses therefore not only need to be vigilant in eliciting and evaluating consumer expectations but also need to be diligent in explaining their role and informing the public of their qualifications.

MSHAKE. A tool for measuring staff knowledge related to geriatric mental health.

This article describes the development of a knowledge-screening instrument used with staff caring for elderly patients with neuropsychiatric disability. A 25-item tool, referred to as the Mary Starke Harper Aging Knowledge Exam (MSHAKE), was developed to verify knowledge competencies of 171 employees at a geropsychiatric center in the Southeastern United States. The MSHAKE assesses basic geriatric mental health knowledge in an educationally diverse health care work force. A variety of descriptive and inferential statistics provide preliminary evidence supporting the use of the MSHAKE as an effective measure of staff members' essential knowledge of aging and neuropsychiatric disorders. This tool has practical application in the clinical setting as part of a competency verification system or in-service evaluation.

The serendipity of faculty practice: strategies for success.

The contemporary social issues impacting healthcare coupled with the increasing demands for academic units to generate income have contributed to the emergence of faculty practice as an integral component of the nurse educator's role. As a result, faculty are encouraged increasingly to assume entrepreneurial joint appointments with the service industry. For nurse educators who engage in faculty practice, serendipity occurs when they immerse themselves in situations and emerge from the experience making unexpected discoveries. The author shares practical recommendations and strategies resulting from a successful 9-month faculty practice.

Perceptions of parenting stress and family relations by fathers of children evaluated for organ transplantation.

18 fathers of children evaluated for solid organ or bone marrow transplantation completed measures of parenting stress and family functioning. Comparisons with normative data indicated that these fathers reported less parenting stress, less family conflict, more concern about family finances, and more limitations in family activities. These data highlight the need for family-based assessments in pediatric transplantation.

Nurse case managers' opinions of their role.

Delphi and focus group methods were used to identify nurse case managers' (NCMs') opinions regarding their professional role. This study convened 15 NCMs from a regional medical center in the southeastern United States. Findings present the highest-rated items influencing the NCM role, as well as a definition and model depicting the role's essential elements. In study questionnaires, participants were asked to identify and subsequently rate items in seven categories. The overall highest-rated item was the personal attribute of critical thinking and prioritizing. The second highest-rated response was the critical function of coordinating a multidisciplinary plan of care. Interdisciplinary support and participative decision making were consistently ranked as important organizational factors. In the category of impact on health care outcomes, identifying patient needs and services was highly rated. Participants suggested that NCM assignments should be based on clinical knowledge and expertise, with a focus on prevention. Practice implications, based on the findings, are emphasized.

Ontogeny and plasma-membrane domain localization of amino acid transport system L in rat liver.

Na(+)-independent hepatic transport of branched-chain amino acids occurs via at least two distinct transport processes. System L1, characterized by micromolar Km values, predominates in hepatoma and fetal hepatocytes, whereas System L2, distinguished by Km values in the millimolar range and sensitivity to inhibition by N-ethylmaleimide (NEM), predominates in adult hepatocytes. To determine the plasma-membrane domain localization and ontogeny of System L activity in the rat, we prepared membrane vesicles from the livers of suckling (10 days old) and adult rats enriched for either basolateral (BLMV) or canalicular (CMV) domains. The initial rate of [3H]leucine uptake into BLMV and CMV derived from adult liver was significantly inhibited by the addition of 5 mM NEM; transport into BLMV and CMV derived from 10-day-old rat liver was not affected. Michaelis-Menten kinetic parameters estimated in BLMV derived from adult liver were consistent with System L2 (Km = 2.16 +/- 0.62 mM, Vmax. = 781 +/- 109 pmol/5 s per mg of protein), as were those estimated in adult CMV (Km = 0.83 +/- 0.21 mM, Vmax. = 385 +/- 38 pmol/5 s per mg of protein). Conversely, kinetic parameters estimated in BLMV derived from livers of suckling rats were consistent with System L1 (Km = 0.041 +/- 0.024 mM, Vmax. = 8.8 +/- 1.5 pmol/5 s per mg of protein), as were those from CMV of suckling rats (Km = 0.023 +/- 0.09 mM, Vmax. = 28.1 +/- 2.1 pmol/5 s per mg of protein). We conclude that NEM-inhibitable Na(+)-independent leucine transport activity consistent with System L2 is present in both BLMV and CMV derived from adult rat liver, whereas System L1 predominates in 10-day-old rat liver tissue.

Taurocholate transport by basolateral plasma membrane vesicles isolated from human liver.

Transport of taurocholate into the hepatocyte against unfavorable chemical and electrical gradients occurs via a sodium-dependent, carrier-mediated transport system. Although this cotransporter has been characterized in the rodent, it has not been demonstrated in man. Therefore, we utilized human liver, obtained via multiorgan donation but not used for transplantation, to prepare basolateral (sinusoidal) liver plasma membrane vesicles by a Percoll gradient method. Na+,K+-ATPase, a marker enzyme for the basolateral domain, was enriched 28.9-fold in the final membrane fraction compared with homogenate, whereas the bile canalicular membrane enzymes Mg++-ATPase and alkaline phosphatase were enriched only 3.4- and 6.4-fold, respectively. Marker enzyme activities for endoplasmic reticulum, lysosomes and mitochondria were not enriched compared with homogenate. Integrity of the membrane vesicles was confirmed by the demonstration of Na+-dependent concentrative uptake of the amino acid L-alanine (estimated intravesicular volume of 0.59 microliter per mg protein). An inwardly directed 100 mM Na+ gradient stimulated the initial rate of 2.5 microM taurocholate uptake and energized a transient 2-fold accumulation of the bile acid above equilibrium ("overshoot"). In contrast, uptake was slower and no overshoot occurred with a K+ gradient. A negative intravesicular potential, created by altering accompanying anions or by valinomycin-induced K+ diffusion potentials, did not enhance taurocholate uptake, suggesting an electroneutral cotransport mechanism. Chloride as the accompanying anion stimulated the initial rate of uptake compared with anions of lesser or greater lipid permeability. Na+-dependent taurocholate (4 microM) uptake was significantly inhibited by 250 microM cholate, taurocholate, glycocholate, taurochenodeoxycholate and bromsulfophthalein.(ABSTRACT TRUNCATED AT 250 WORDS)

Recent advances in mammalian amino acid transport.

During the last four decades, mammalian amino acid transport systems have been described at the cellular level through general properties such as ion-dependence, kinetics, substrate specificity, regulation of activity, and numerous other characteristics. These studies have allowed the definition of multiple transport systems for neutral, anionic, and cationic amino acids. Each system is distinct but exhibits overlapping substrate specificity. Direct measurement of transport has permitted a wealth of information to be accumulated regarding the regulation of overall activity, but the underlying molecular mechanisms have not been investigated because of a lack of the appropriate tools. Recent research designed to obtain these tools has proven fruitful, and the field of amino acid transport clearly is entering a new era. In the immediate future, transporter properties such as hormonal regulation, adaptive control, ion-dependence, and trans-effects will be studied at the molecular level by assaying mRNA or protein content and by analyzing results obtained with altered protein structures following site-directed mutagenesis. Identification of specific proteins associated with activities already well described will provide answers to heretofore untestable questions. For example, is Na(+)-independent transport mediated by the same proteins that mediate Na(+)-dependent uptake except that their function in this mode does not require sodium binding? What is the protein composition of amino acid transporters? As discussed above, emerging evidence suggests that transporter proteins have different molecular structure, 12 versus 1 transmembrane domains, or that they exist as heterodimers or heterotetramers. Identification of certain transporter proteins and cloning of the respective genes also will provide valuable information about a number of inheritable diseases that are thought to be caused by defects in transporter synthesis or function. The opportunity to ask these questions will certainly generate renewed interest in the field of amino acid transport and lead to exciting advances in our knowledge.