Alterations of surfactant pools in fetal and newborn rat lungs

Preparation by the developing alveolar epithelium for the transition to air breathing and surfactant secretion at birth are critical components of neonatal survival. We combined morphometric analysis and biochemical assays of lung phospholipids to measure the amount and redistribution of lung surfactant during the perinatal period of rats. Within 10 min of the start of air breathing, there was a small increase in type II cell lamellar body content by morphometric and biochemical estimates. By 24 h, the whole lung and alveolar extracellular pool surfactant lipid had substantially increased. Subfractionation of the alveolar surfactant pool obtained at four times, from birth to 24 h of life, demonstrated a 20-fold increase in the ratio of phospholipid in a tubular myelin-rich fraction compared to a unilamellar vesicle-rich fraction. We conclude that packaging of surfactant may be very active immediately postbirth. Our results also indicate a major shift in the physical forms of extracellular surfactant during the first hours of air breathing.     

Exogenous cholecystokinin (CCK) reduces neonatal rat brain opioid receptor density and CCK levels

Newborn rats were given saline or cholecystokinin8 (CCK8) (5 micrograms/kg, twice daily) i.p. for 3 weeks. On day 21, effects on brain development were assessed. CCK-like immunoreactivity was measured in 7 brain regions; a small (12-18%) but significant decrease in endogenous levels of this peptide was detected in cerebral cortex, medulla and pons of the CCK-treated rats. Morphometric measurements revealed a slight reduction in thickness of most cerebral cortical sections within the CCK-treated group. The area of a midsagittal section of the cerebellum was unchanged except for the Purkinje/granule cell layer, which was smaller in CCK-treated animals. Levels of mu-, delta- and kappa-opioid receptors were estimated by homologous displacement binding assays using selective radioligands. The CCK treatment resulted in a significant decrease in levels of mu- (11%) and delta- (13%)-sites in the cerebral cortex. Neither binding affinities nor kappa-receptor densities were altered. Other animals received the same treatment regimens for 21 days and were maintained for an additional 29 days without treatment; these rats had reductions only in cortical mu-sites (15%). Chronic intraventricular administration of CCK (0.1 microgram/h) to adult rats did not elicit a similar down-regulation of cortical mu or delta receptors, suggesting that the effects observed in neonates reflected developmental processes.     

Influence of anomalous rectifier activation on afterhyperpolarizations of neurons from cat sensorimotor cortex in vitro

1. Large neurons from layer V of cat sensorimotor cortex (Betz cells) were studied to determine the influence of the anomalous rectifier current (IAR) on slow afterhyperpolarizations (AHPs). The neurons were examined using intracellular recording and single-microelectrode voltage clamp in an in vitro brain slice preparation. 2. A faster medium-duration AHP (mAHP) and slower AHP (sAHP) followed repetitive firing (22, 23). The amplitude of the mAHP often increased or remained constant during membrane potential hyperpolarization. The membrane potential trajectory resulting solely from IAR activation was similar to the mAHP. 3. Postrepetitive firing voltage clamp was used to measure directly slowly decaying K+ currents (IK) and IAR at different membrane potentials. IK exhibited both a fast and slow decay. The time constants of the fast decay of IK and IAR activation were similar. IAR increased with hyperpolarization or raised extracellular K+ concentration [( K+]o), whereas both the fast and slow components of IK reversed or nulled near -100 mV and behaved as pure K+ currents in response to raised [K+]o. 4. To determine the precise contribution of IK and IAR to the AHP waveform, theoretical AHPs were computed using a quantitative model based on voltage-clamp measurements. The calculated AHPs were qualitatively similar to measured AHPs. The amplitude of the mAHP showed little change with hyperpolarization because of the increasing dominance of IAR at more negative membrane potentials. The sAHP was little affected by IAR activation. 5. Several model parameters subject to biological variation among Betz cells were varied in the calculations to determine their importance in the AHP waveform. With IK parameters held constant, the amplitude and time course of the mAHP depended on resting potential, membrane time constant, the kinetics of the anomalous rectifier conductance (GAR), and the maximum value of GAR. IAR activation could result in a biphasic AHP even when the fast decay of IK was omitted from the calculations. 6. A wider variation of model parameters revealed behavior that may be relevant to other neurons. Certain values of membrane or IAR activation time constants resulted in a monophasic AHP even when the fast decay of IK was present. The decay of a biphasic AHP could reflect either the onset of IAR or the fast decay of IK, depending on the relative value of their time constants. Procedures are outlined to discriminate between these possibilities using current clamp methods.(ABSTRACT TRUNCATED AT 400 WORDS)     

Slow conductances in neurons from cat sensorimotor cortex in vitro and their role in slow excitability changes

1. The electrophysiological and pharmacological properties of slow afterpotentials in large layer V neurons from cat sensorimotor cortex were studied in an in vitro slice preparation using intracellular recording and single-microelectrode voltage clamp. These properties were used to assess the role of afterpotential mechanisms in prolonged excitability changes. 2. The mean duration of a slow afterhyperpolarization (sAHP) was 13.5 s following 100 spikes evoked at 100 Hz. Its time course was best described by two exponential components, which decayed with time constants of several hundred milliseconds (the early sAHP) and several seconds (the late sAHP). The amplitude of both the early and late components were sensitive to membrane potential and raised extracellular K+ concentration [( K+]o). 3. The early sAHP was reduced when divalent cations were substituted for Ca2+, whereas the late sAHP was unaffected. We conclude that a Ca2+-mediated K+ conductance is responsible for much of the early sAHP. In the presence of tetrodotoxin (TTX), 1-s voltage-clamp steps were used to evoke slow AHPs or outward ionic currents. These AHPs and currents were abolished in Ca2+-free perfusate, but they had a maximum duration of only a few seconds. Thus the slowest outward currents we could observe during voltage clamp in TTX were responsible only for the early sAHP. 4. The possible role of an electrogenic Na+-K+ pump in the late sAHP was examined by applying ouabain to the slice. Ouabain did not reduce selectively the late sAHP, and its effect was best explained by a decrease in intracellular K+ concentration and an increase in [K+]o. 5. Muscarinic and beta-adrenergic agonists reduced or abolished the entire (early and late) sAHP. Neither type of agonist affected the Ca2+-dependent, apamin-sensitive medium-duration afterhyperpolarization (35). We conclude that both the Ca2+-mediated K+ conductance underlying the early sAHP and the Ca2+-independent mechanisms underlying the late sAHP are sensitive to at least two classes of transmitter agonists. 6. We focused on the muscarinic effects. When concentrations greater than 5 microM were employed, the entire (early and late) sAHP was replaced by a slow afterdepolarization (sADP). Muscarine reduced the sAHP directly by reducing the underlying outward ionic currents and indirectly by causing the sADP. The sADP was Ca2+-mediated, since it was abolished by Ca2+-free perfusate but not by TTX. 7. The ionic currents underlying the sAHP and the sADP influenced excitability for seconds following evoked repetitive firing.(ABSTRACT TRUNCATED AT 400 WORDS)     

Detection and characterization of beta-adrenergic receptors and adenylate cyclase in coated vesicles isolated from bovine brain

To assess whether internalization of beta-adrenergic receptor occurs in the CNS, we have isolated clathrin-coated vesicles from bovine forebrain and examined them for the presence of beta-adrenergic receptor binding and adenylate cyclase activities. A coated vesicle enriched preparation isolated by successive D2O-Ficoll density gradient centrifugations was applied to a glass bead permeation column to achieve further purification. Two major peaks of protein were eluted from the column and monitored by electron microscopy and SDS-PAGE. Peak II contained almost exclusively coated vesicles (98%), whereas peak I, which appeared in the void volume, contained larger smooth vesicles and few coated vesicles. beta-Adrenergic receptor binding to peaks I and II was measured with 125I-cyanopindolol (CYP) as ligand in Sepharose 4B column assays. 125I-CYP was found to bind specifically and saturably to both peaks I and II with a Bmax of 28 +/- 4 and 32 +/- 3 fmol/mg protein, respectively. 3H-CGP 12177, a hydrophilic beta-adrenergic receptor ligand, did not label receptors present in peak II, but it specifically bound to synaptic plasma membranes (SPM) prepared from bovine hippocampus and, to a lesser extent, to peak I. These results suggest that receptors present in coated vesicles are cryptic in nature. In the displacement of 125I-CYP binding by (-)-isoproterenol, addition of 50 microM GppNHp caused a significant "right shift" with SPM and peak I but not the peak II preparation. Adenylate cyclase activities could also be detected in both peaks I and II (specific activities, 21 +/- 0.6 and 24 +/- 0.5 pmol cAMP/mg protein/min, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Pulmonary contusions in patients with rib fractures: The need to better classify a common injury

BackgroundPulmonary contusions are common injuries. Computed tomography reveals vast contused lung volume spectrum, yet pulmonary contusions are defined dichotomously (unilateral vs bilateral). We assessed whether there is stepwise increased risk of pulmonary complications among patients without, with unilateral, and with bilateral pulmonary contusion.MethodsWe identified adults admitted with rib fractures using the largest US inpatient database. After propensity-score-matching patients without vs with unilateral vs bilateral pulmonary contusions and adjusting for residual confounders, we compared risk for pneumonia, ventilator-associated pneumonia (VAP), respiratory failure, intubation, and mortality.ResultsAmong 148,140 encounters of adults with multiple rib fractures, 19% had concomitant pulmonary contusions. Matched patients with pulmonary contusions had increased risk of pneumonia 19% [95%CI:16–33%], respiratory failure 40% [95%CI: 31–50%], and intubation 46% [95%CI: 33–61%]. Delineation showed bilateral contusions, not unilateral contusions, attributed to increased risk of complications.ConclusionsThere is likely a correlation between contused lung volume and risk of pulmonary complications; dichotomously classifying pulmonary contusions is insufficient. Better understanding this correlation requires establishing the clinically significant contusion volume and a correspondingly refined classification system.