Effects of hypoxia on membrane potential and intracellular calcium in rat neonatal carotid body type I cells

1. We have studied the effects of hypoxia on membrane potential and [Ca2+]i in enzymically isolated type I cells of the neonatal rat carotid body (the principal respiratory O2 chemosensor). Isolated cells were maintained in short term culture (3-36 h) before use. [Ca2+]i was measured using the Ca(2+)-sensitive fluoroprobe indo-1. Indo-1 was loaded into cells using the esterified form indo-1 AM. Membrane potential was measured (and clamped) in single isolated type I cells using the perforated-patch (amphotericin B) whole-cell recording technique. 2. Graded reductions in PO2 from 160 Torr to 38, 19, 8, 5 and 0 Torr induced a graded rise of [Ca2+]i in both single and clumps of type I cells. 3. The rise of [Ca2+]i in response to anoxia was 98% inhibited by removal of external Ca2+ (+1 mM EGTA), indicating the probable involvement of Ca2+ influx from the external medium in mediating the anoxic [Ca2+]i response. 4. The L-type Ca2+ channel antagonist nicardipine (10 microM) inhibited the anoxic [Ca2+]i response by 67%, and the non-selective Ca2+ channel antagonist Ni2+ (2 mM) inhibited the response by 77%. 5. Under voltage recording conditions, anoxia induced a reversible membrane depolarization (or receptor potential) accompanied, in many cases, by trains of action potentials. These electrical events were coincident with a rapid rise of [Ca2+]i. When cells were voltage clamped close to their resting potential (-40 to -60 mV), the [Ca2+]i response to anoxia was greatly reduced and its onset was much slower.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development of glutamate-induced intracellular Ca2+ rise in the embryonic chick retina

Plasmids derived from bacteriophage lambda are known as lambda plasmids. These plasmids contain the ori lambda region and lambda replication genes O and P. Typical lambda plasmids also contain the cro gene, the product of which is a repressor of the pR promoter when present at relatively high concentrations. These genes stably maintain the plasmid in Escherichia coli at copy numbers of 20 to 50 per cell. According to a generally accepted model, stable maintenance of lambda plasmids is possible due to the Cro repressor autoregulatory loop (the cro gene is under control of pR). Here we demonstrate that lambda plasmids devoid of the Cro autoregulatory loop can also be stably maintained in E. coli strains. We present data for two such plasmids: pTC lambda 1 in which the pR-cro region has been replaced by the ptetA promoter and the tetR gene (coding for the TetR repressor), and a standard lambda plasmid with inactivated cro gene (lambda cro-null plasmid). Thus, the presence of the Cro repressor autoregulatory loop does not appear to be essential to the maintenance of lambda plasmids in vivo.     

Changes in Bruch's membrane and related structures with age

Age-related macular disease is a major and growing public health burden in developed Caucasian societies, accounting for about 50% of blind registration. Evidence exists that this is an emerging problem in Eastern Asia, although the phenotype appears to differ from that seen in Western society. It is likely that several genes are involved, and that the genes or allelic variants conferring are common. Environment plays a major role in its pathogenesis, and it is believed that genetic susceptibility becomes apparent only if there are sufficient environmental pressures. There is no therapy currently available that will have an impact on the prevalence of blindness from age-related macular disease. It has been shown that visual loss occurs as a reaction to ageing changes in Bruch's membrane, which is interposed between the choriocapillaris and the retinal pigment epithelium. The age changes in all three structures have been partly characterised, and as a consequence, multiple putative pathogenic mechanisms have been proposed. Cross-sectional studies of populations with different genetic background and life styles would serve to prove the importance of inheritance and environment. Molecular genetic analysis of blood from affected sibling pairs from these sources may indicate the relevant genes, the prevalence of which may differ in different communities. Enquiries as to life styles may determine important environmental influences. Examination of donor eyes from these communities may reveal distinctive features that may reflect the variation in genetic predisposition and environmental pressures. It is hoped that the findings from such studies will lead to novel and potentially successful management strategies.     

Na+-H+ exchange and intracellular pH regulation in colonocytes from the chick

The nutritional effects on operant visual learning behavior were investigated in a flight simulator. Operant visual learning and memory formation were normal in Drosophila (S-flies) reared on standard medium, but absent in flies (P-flies) raised on Peking medium. S- and P-flies were transferred to the alternative medium soon after hatching and their progeny was also raised on corresponding medium for several generations. respectively. S-flies transferred showed significantly reduced learning acquisition and 20 min memory retention, and operant visual learning along with memory formation was abolished in their progeny within three generations. Transferred P-flies recovered slowly their learning acquisition and memory formation to normal levels within five generations. Further studies suggested that low protein and minerals or high carbohydrate contents in Peking medium might be related to abnormal performance of P-flies. These results confirm the feasibility of affecting learning behavior by dietary regimens and developing an insect model of maternal malnutrition for pre- or post-natal malnutrition in Drosophila.     

Potentiometric study of resting potential, contributing K+ channels and the onset of Na+ channel excitability in embryonic rat cortical cells

Resting membrane potential (RMP), K+ channel contribution to RMP and the development of excitability were investigated in the entire population of acutely dissociated embryonic (E) rat cortical cells over E11-22 using a voltage-sensitive fluorescent indicator dye and flow cytometry. During the period of intense proliferation (E11-13), two cell subpopulations with distinct estimated RMPs were recorded: one polarized at approximately -70 mV and the other relatively less-polarized at approximately -40 mV. Ca2+o was critical in sustaining the RMP of the majority of less-polarized cells, while the well-polarized cells were characterized by membrane potentials exhibiting a approximately Nernstian relationship between RMP and [K+]o. Analysis of these two subpopulations revealed that > 80% of less-polarized cells were proliferative, while > 90% of well-polarized cells were postmitotic. Throughout embryonic development, the disappearance of Ca2+o-sensitive, less-polarized cells correlated with the disappearance of the proliferating population, while the appearance of the K+o-sensitive, well-polarized population correlated with the appearance of terminally postmitotic neurons, immuno-identified as BrdU-, tetanus toxin+ cells. Differentiating neurons were estimated to contain increased K+i relative to less-polarized cells, coinciding with the developmental expression of Cs+/Ba2+-sensitive and Ca2+-dependent K+ channels. Both K+ channels contributed to the RMP of well-polarized cells, which became more negative toward the end of neurogenesis. Depolarizing effects of veratridine, first observed at E11, progressively changed from Ca2+o-dependent and tetrodotoxin-insensitive to Na+o-dependent and tetrodotoxin-sensitive response by E18. The results reveal a dynamic development of RMP, contributing K+ channels and voltage-dependent Na+ channels in the developing cortex as it transforms from proliferative to primarily differentiating tissue.     

Effects of stimulation on embryonic activity in the chick.

Studied the effects of stimulation on the stereotyped and coordinated movements in the chick (Rhode Island Red/Light Sussex hybrids) embryo using the type of stimulation (regular loud clicks) known to accelerate development and the time of hatching. The number of coordinated movements, the amount and approximate size of all activity, heart rates and, in 20-day embryos, respiration rates were assessed during 15-min control, stimulation, and no stimulation periods. Results show an increase in the number of coordinated movements, but not their size, and some changes in the heart and respiration rates after the onset of stimulation. Effects occurred mostly after several minutes' delay. It is suggested that this delay may be connected with the reciprocal relationship between the coordinated movements and random motility in that the former may be triggered more readily at times when the latter are suppressed. (20 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Role of the Na+/K+-ATPase in regulating the membrane potential in rat peritoneal mast cells

500 children with ages ranging between 2 and 13 years and their families joined the study. A detailed questionnaire and a 3-day food intake diary were evaluated to find the relationship between dental caries and dietary habits. It is concluded that the prevalence of caries increases by frequent and high sugar consumption. With lower age and lower caries experience followed a more balanced diet. There was an inverse relationship between caries prevalence and the mothers' educational level. The percentage of children who never or irregularly brushed their teeth was highest in the caries active group.     

Androgens and imprinting: Differential effects of testosterone on filial preference in the domestic chick.

After hatching, chicks received either a subcutaneous injection of testosterone enanthate (0.5–5 mg) in oil or oil alone (controls). Ss were then trained by exposing them to a rotating imprinting stimulus, either a red box or a stuffed jungle fowl. A preference score was then determined, providing a measure of the strength of imprinting. Plasma testosterone concentration after testing did not differ significantly between males and females and was unaffected by the type of training stimulus. In fowl-trained Ss, preference score was positively correlated with plasma testosterone concentration. The mean preference score of the fowl-trained Ss that had received exogenous testosterone was significantly higher than that of controls; no such effect was found in box-trained Ss. A previous study by D. C. Davies et al (1985) showed that imprinting with the box, but not the fowl, was profoundly impaired by a noradrenergic neurotoxin. Results suggest that some of the neural systems supporting the preference for a simple artificial object are different from some of those supporting the preference for the stuffed jungle fowl. (23 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Differential effects of age on involuntary and voluntary autobiographical memory.

Research on aging and autobiographical memory has focused almost exclusively on voluntary autobiographical memory. However, in everyday life, autobiographical memories often come to mind spontaneously without deliberate attempt to retrieve anything. In the present study, diary and word-cue methods were used to compare the involuntary and voluntary memories of 44 young and 38 older adults. The results showed that older adults reported fewer involuntary and voluntary memories than did younger adults. Additionally, the life span distribution of involuntary and voluntary memories did not differ in young adults (a clear recency effect) or in older adults (a recency effect and a reminiscence bump). Despite these similarities between involuntary and voluntary memories, there were also important differences in terms of the effects of age on some memory characteristics. Thus, older adults’ voluntary memories were less specific and were recalled more slowly than those of young adults, but there were no reliable age differences in the specificity of involuntary memories. Moreover, older adults rated their involuntary memories as more positive than did young adults, but this positivity effect was not found for voluntary memories. Theoretical implications of these findings for research on autobiographical memory and cognitive aging are discussed. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

A primary role for K+ and Na+ efflux in the activation of apoptosis

Cell shrinkage is a major characteristic of apoptosis, but the mechanism and role of this process in cell death are poorly understood. The primary factor that controls volume regulation in all cells is ions, and thus we have examined the movement of ions at the single cell level in lymphocytes during apoptosis. Activation of the death program with several stimuli that act through independent pathways to stimulate apoptosis results in a synchronous shift of cells from a normal cell size to a shrunken cell size. Only the shrunken cells exhibit DNA fragmentation and an approximate 4-fold elevation of caspase-3-like activity. Analysis of K+ and Na+ ion content of individual cells by flow cytometry revealed that the intracellular ionic strength of apoptotic cells decreased substantially from their non-shrunken counterparts. Additionally, we show apoptosis is enhanced under conditions where the intracellular K+ concentration is diminished and that apoptosis is inhibited when K+ efflux is prevented. These data show that the efflux of ions, primarily potassium, plays a necessary and perhaps a pivotal role in the cell death program.     

Changing activation sequence in the embryonic chick heart. Implications for the development of the His-Purkinje system

Viruses that establish persistent infections in their host, such as herpesviruses, adenoviruses or HIV, express proteins designed to pre-empt or evade recognition and elimination by MHC class I restricted CD8+ T lymphocytes. Notable discoveries during the annual period of review have demonstrated that, in principle, each single step within the MHC class I pathway of antigen processing and presentation is fair game for manipulation by viral functions. The viral factors that are natural inhibitors of this pathway have been instrumental for the elucidation of the distinct molecular mechanisms that are exploited by viruses. The viral stealth strategies that downregulate MHC class I protein surface expression may lead, however, to a higher susceptibility of virus-infected cells to natural killer cell activity. Strikingly, there is evidence that some viruses counteract increased natural killer cell recognition by expressing viral MHC class I homologues that function as surrogate inhibitors of natural killer cell activity.     

Differential class III and glibenclamide effects on action potential duration in guinea-pig papillary muscle during normoxia and hypoxia/ischaemia

1. Microelectrode recording techniques were used to study the effects of several potassium channel blockers which are considered to be Class III antiarrhythmic compounds. The effects of (+)-sotalol, UK-66,914, UK-68,798 and E-4031 on action potential duration (APD) were determined in guinea-pig isolated papillary muscles. The compounds were evaluated under normoxic or hypoxic/ischaemic conditions at 36.5 degrees C and compared to glibenclamide, which is considered to be a blocker of ATP-dependent potassium channels. Prolongation of action potential duration at 90% repolarization (APD90) was taken as an indirect measure of potassium channel blockade. 2. Under normoxic conditions, the Class III compounds prolonged APD in a concentration-dependent manner. According to EC15 values, the order of potency of the Class III compounds was found to be UK-68,798 > E-4031 > UK-66,914 > (+)-sotalol. Glibenclamide did not significantly prolong APD90 under normoxic conditions. 3. Perfusion with an experimental hypoxic or ischaemic bathing solution produced qualitatively similar effects on action potentials. Over a period of 20-25 min in either of the experimental solutions, there was a small decrease in action potential amplitude (APA) and a prominent shortening of APD. The ischaemic solution also depolarized the resting membrane potential by about 15 mV. 4. (+)-Sotalol and UK-66,914 did not reverse the shortening of APD induced by perfusion with hypoxic Krebs solution. High concentrations of glibenclamide (10 microM) and UK-68,798 (30 and 60 microM) partially reversed the hypoxia-shortened APD. Glibenclamide was more potent and exhibited a greater time-dependent action than UK-68,798. 5. During experimental ischaemia, the Class III compound E-4031 (10 microM, n = 7) produced small, but significant, increases in the APD90 (11 +/-3 ms after 20 min) which were not clearly time-dependent(14 +/- 4 ms after 30 min). UK-68,798 (10 microM) also produced a small, but insignificant, increase in APD90(12 =/-6 ms at 20 min, n = 4). Higher concentrations of UK-68,798 (30 and 60 microM, n = 4) did not produce a consistently significant increase in APD90 during ischaemia: significance was only attained after 20 min in the presence of 60 microM UK-68,798 (24 +/- 12 ms). However, in marked contrast to the effects of the Class III compounds, glibenclamide (10 microM) produced large time-dependent increases in ischaemic APD90 (34 +/- 11 ms at 7 min, n = 9) which were significant 15 min or more after drug addition(52 +/- 12 ms at 20 min, n = 7; 74 +/- 5 ms at 30 min, n = 6).6. The present microelectrode data suggest that blockers of ATP-dependent potassium channels, such as glibenclamide, might prove to be more effective than Class III compounds against ischaemia-induced shortening of cardiac action potentials.     

Role of the steady-state Na+ channel current in pacemaker depolarizations in young embryonic chick ventricular myocytes

Thresholds to noxious mechanical and thermal stimulation were measured in 6 groups of sheep prior to induction of anaesthesia and subsequently for a period of 2 h in the post-anaesthetic period. Groups 1-4 were anaesthetised using thiopentone and underwent ventral midline laparotomy. Four animals (group 5) underwent anaesthesia but not surgery, and a further 6 sheep (group 6) undergoing surgery were anaesthetised using ketamine. Groups 1-3 were intravenously administered the following drugs intra-operatively: flunixin meglumine, carprofen and buprenorphine, respectively. Groups 4-6 received no additional treatment. Thresholds to the mechanical test were not changed in the post-anaesthetic period for any group. There was a significant reduction in the responses to thermal stimulation after surgery for sheep in group 4 (45 and 60 min), while sheep in group 2 had thresholds to thermal stimulation greater than those recorded in the remaining groups at all time points post-operatively. Responses to thermal stimulation in sheep undergoing anaesthesia but not surgery (group 5) were unaltered during the 2 h recording period after anaesthesia ended. These data indicate that abdominal surgery induces thermal but not mechanical hyperalgesia in sheep, which appears to be centrally mediated. Moreover, the absence of mechanical hyperalgesia raises the possibility that central changes in noxious information processing may not be detected using mechanical stimuli in the same time course as thermal stimuli.     

Effect of concanavaline A on intracellular K+ and Na+ concentration and K+ transport of human lymphocytes

Incubation of human lymphocytes with ConA causes an increase in [Na+]i and a decrease in [K+]i. This effect is not due to the experimental washing procedure, but is due to the ConA-induced increase in permeability which is not fully compensated by the increase in active transport. The ConA-induced increase in 42K+ uptake consists of an increase in leak flux which is independent of [Na+]o, and of an increase in pump flux which is dependent on [Na+]o. The increase in leak flux may be caused by increased membrane fluidity. The increase in pump flux may be produced by the increased [Na+]i and by a stimulation of Na+, K+ATPase.     

Differential binding activity of erythrocyte ankyrin to the alpha-subunits of Na+, K(+)-ATPases from rat cerebral and axonal membrane

Pertussis toxin catalysed the ADP-ribosylation of a protein of M(r) 40,000 in ovine luteal tissue. Ribosylation of 45% of this protein in whole cell incubations (as judged by subsequent ribosylation of cell-free preparations in the presence of [32P]NAD) attenuated the prostaglandin (PG)F2 alpha-stimulated hydrolysis of [3H]inositol-labelled phosphatidylinositol-4,5-bisphosphate into inositol trisphosphate by 60%, but did not affect the inhibition by PGF2 alpha of LH-stimulated accumulation of cyclic AMP. It is concluded that activation of phospholipase C by PGF2 alpha involves a pertussis toxin-sensitive protein, probably a G protein, and that the inhibitory effect of PGF2 alpha on LH-stimulated adenylate cyclase is unlikely to be directly mediated by such a protein.     

Differential effects of indomethacin on the sheep ovary: prostaglandin biosynthesis, intracellular calcium, apoptosis, and ovulation

Cells of the apical wall of the dominant follicle and contiguous ovarian surface epithelium become apoptotic with the approach of ovulation in the sheep. It was hypothesized that indomethacin, an established inhibitor of prostaglandin biosynthesis and ovulation, would protect apical ovarian cells from programmed death. The anovulatory potencies of two systemic doses of indomethacin (200 and 800 mg) were tested in gonadotropin-stimulated ewes. A complete blockade of ovulation occurred at the higher dose of indomethacin. Ovulation was not inhibited by 200 mg indomethacin. Both doses of drug suppressed follicular prostaglandin production below pregonadotropin levels. Immunofluorescence detection of digoxigenin end-labeled (fragmented) DNA was used as a marker of apoptosis among ovarian surface epithelial and granulosa cells recovered from the optical hemisphere of preovulatory ovine follicles. Cellular DNA fragmentation was averted in animals given 800 mg indomethacin, whereas apoptosis ensued after 200 mg. A sustained increase in cytosolic calcium is generally a prerequisite to apoptotic DNA fragmentation and cell death. Indeed, intracellular calcium, detected by fluorescence of fura-2, was elevated in ovarian cells of animals destined to ovulate (controls, 200 mg indomethacin) in comparison to (safeguarded) cells of anovulatory ewes (800 mg indomethacin). These observations provide circumstantial evidence that apical ovarian cell degeneration by calcium-mediated apoptosis is a determinant of follicular instability and rupture, but that these events are unrelated to the gonadotropin-induced rise in prostanoid production characteristic of preovulatory follicles.     

Cardiovascular effects of a new inotropic agent, U. K. 14275, in patients with coronary heart disease

1 The chronotropic and inotropic properties of U.K. 14275, a phosphodiesterase inhibitor were assessed in patients with coronary heart disease. 2 Left ventricular function was assessed in eight patients with acute myocardial infarction using the non-invasive measurement of systolic time intervals. 3 Twelve patients with angina pectoris were studied during diagnostic coronary arteriography. Left ventricular function was assessed using a high fidelity catheter tipped transducer in the left ventricle. 4 In both groups of patients U.K. 14275 infused intravenously in doses of 32, 64, 128 and 256 microgram kg-1 bodyweight min-1 enhanced the contractile state of the left ventricle without altering the heart rate.     

The effects of administering lithium carbonate on the balance of Na, K and water in manic-depressive patients

Eleven patients in remission from manic-depressive illness were studied by means of metabolic balances before and after the administration of lithium carbonate. Lithium caused a sharp diuresis of isotonic saline and a smaller excretion of potassium over the course of two days. During the subsequent two days there occurred a compensatory retention of Na, K and water. These short-term changes were not associated with any significant alteration in the patients' mood. There was no significant and systematic retention of Na, K or water over the 14 days of Li administration. The recovery of Li was measured simultaneously. During the first week only a proportion of the administered Li was recovered in the urine and faeces, suggesting that a gradual distribution of Li throughout its body space was occurring. After the first week, nearly all the administered Li was recovered, indicating an equilibrium with an even distribution of the ion throughout its body space. This equilibrium was more complete at this early stage in those patients who had been given a smaller dose of lithium carbonate.