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PubMed1986

Circadian, sleep and brain temperature rhythms in cats under sustained daily light-dark cycles and constant darkness.

In adult cats, sleep-wake states and brain temperatures were determined under experimental LD cycles and prolonged DD. Convincing rhythms had not previously been well-established for this animal. The present results demonstrated daily fluctuation in total sleep time (TST) and in brain temperature (Tb). These comprise predominantly nocturnal wakefulness and high temperatures but include a bi-modal component marking dawn and dusk peaks. In addition there are ultradian variations with a period of 2-4 hour superimposed on the circadian rhythm. The latter continued at the beginning of prolonged DD, but gradually diminished and disappeared after about ten days. The TST circadian rhythm diminished more quickly than the Tb rhythm. This result suggests that the former is coupled more strongly to the circadian oscillator than the latter, and that the brain temperature rhythm could not depend on the circadian rhythm of sleep-wakefulness. These results encourage use of cats in further chronobiological studies of sleep and other physiological rhythms.

Activity Cycles↗
PubMed1984

Protons suppress the dark current of frog retinal rods.

Outer segments of rod photoreceptors with the attached ellipsoid region of the inner segment were isolated from Rana pipiens retinae, and their membrane photocurrents measured with the suction electrode technique in the 'ellipsoid-in' configuration. Under dark adapted conditions in standard Ringer solution, isolated rod outer segments with ellipsoids exhibited maximal photocurrents of 10-30 pA, and light sensitivities of 0.2-1.0 pA/isomerization. A local perfusion technique was employed to change rapidly the solution bathing the outer segment. Rods were tested for their sensitivity to protons by perfusion with Ringer solution of altered pH. The dark current was reversibly suppressed by low pH: in Ringer solution with Calcium activity aCa = 10(-3)M dark current suppression obeyed a hyperbolic saturation law with apparent dissociation constant, pKa = 4.8. The decay of dark current of rods following poisoning with ouabain was retarded by low pH perfusion, as it was by light. Protons thus act to suppress the outer segment Na+ conductance. Three experiments support the hypothesis that protons act interior to the plasma membrane in suppressing the dark current. (1) Perfusion of rods at constant pH with Ringer solution having increased CO2 suppressed the dark current. (2) Removal of perfusate containing 50 mM-NH4Cl causes transient dark current suppression. (3) Acetate, which acts as a neutral proton carrier, when added to Ringer solution, shifts the apparent pKa of dark current suppression to a higher pH. Dark current suppression by protons and recovery occurred with a time constant of ca. 1 s. Low pH perfusion retarded the recovery of the dark current from a saturating flash, slowed the light response in its linear range, and increased light sensitivity. Perfusion at pH = 10.5 caused a slight increase in dark current, sped up the recovery of the rod from a saturating flash, accelerated the linear response and decreased the light sensitivity. Lowering aCa of the Ringer solution caused the proton sensitivity of the dark current to drop. At aCa = 5 X 10(-6) M the apparent pKa of dark current suppression was shifted about 0.8 pH units to pH = 4.0. Cells at aCa = 10(-9) M were insensitive to pH = 3.5, which completely suppressed the dark current at normal aCa. Lowered aCa decreased light sensitivity. Both proton sensitivity and light sensitivity of dark current suppression were estimated for each member of a population of rods in various aCaS:proton sensitivity and light sensitivity were found to be linearly correlated over a range of 3 log units.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed2006

Darkness causes myopia in visually experienced tree shrews.

PURPOSE: To examine the effect of a period of continuous darkness on the refractive state and vitreous chamber depth of normal light-reared juvenile tree shrew eyes, and to learn whether eyes that developed myopia in response to monocular minus-lens wear will recover in darkness. METHODS: Starting at 16 days of visual experience (VE), the refractive state of five dark-treatment tree shrews was measured daily to confirm that it was stable and nearly emmetropic. After corneal and ocular component dimension measures, the animals were placed into continuous darkness for 10 days. On removal of the animals from darkness, corneal and ocular component measures were repeated, and daily refractive measures were resumed. The refractive state of the dark-treatment group was compared with that of a normal-lighting group (n = 5) that received standard colony lighting throughout the measurement period. Five dark-recovery animals wore a monocular -5-D lens for 11 days to induce myopia before they were placed into continuous darkness for 10 days. RESULTS: The animals in the normal-lighting group completed the emmetropization process, stabilizing at approximately (mean +/- SEM) 0.7 +/- 0.3 D of hyperopia (noncycloplegic refraction, corrected for the small eye artifact) at 60 days of VE. Dark-treatment group eyes shifted toward myopia (mean +/- SEM, -4.3 +/- 0.5 D) in the dark. The vitreous chamber became elongated by 0.09 +/- 0.02 mm relative to normal eyes. Corneal power showed a small, near-normal decrease (1.4 +/- 0.3 D). Four of five myopic eyes in the dark-recovery group became more myopic (-2.2 +/- 0.9D) in darkness, and all the fellow control eyes shifted toward myopia (-2.8 +/- 0.5 D). CONCLUSIONS: Maintaining emmetropia is an active process. After eyes have achieved emmetropia or have compensated for a minus lens, continued visual guidance is necessary to maintain a match between the axial length and the focal plane or for recovery to occur. Absence of light is myopiagenic in tree shrews that have developed with normal diurnal lighting. This result contrasts with the apparent absence of a darkness effect in tree shrews reared in the dark from before normal eye opening.

Animals↗
PubMed1996

Vestibular dark cells contain the Na+/H+ exchanger NHE-1 in the basolateral membrane.

Vestibular dark cells and strial marginal cells transport K+ by similar mechanisms. We have shown that K+ transport in vestibular dark cells is sensitive to the cytosolic pH (pHi) (Wangemann et al. (1995a): J. Membrane Biol. 147: 255-262). The present study addresses pharmacologically the questions whether vestibular dark cells and strial marginal cells from the gerbil contain a Na+/H+ exchanger (NHE) and in which membrane, apical or basolateral, NHE is located. Further, the study addresses the question which NHE subtype is present in vestibular dark cells. pHi was measured micro-fluorometrically with the pH-sensitive dye 2',7'-bicarboxyethyl-5(6)-carboxyfluorescein (BCECF), cell volume which is a measure of the net balance between ion influx and efflux was monitored as cell height (CH) and the equivalent short circuit current (Isc) which is a measure of transepithelial K+ secretion was calculated from measurements of the transepithelial voltage (Vt) and the transepithelial resistance (Rt). Changes in pHi were induced by 20 or 40 mM propionate. In the presence of propionate a transient acidification of pHi was observed in vestibular dark cells as well as a subsequent alkalinization to pHi values exceeding those under control conditions. The alkalinization of pHi in the presence of propionate was inhibited by the NHE blockers amiloride and EIPA. Propionate-induced swelling of vestibular dark cells was inhibited by amiloride. The NHE blocker amiloride caused in vestibular dark cells an acidification of pHi and a decrease in CH. Amiloride caused in both vestibular dark cells and strial marginal cells a transient stimulation of Isc when added to the basolateral side but not added to the apical side. Similar effects and pHi were observed in vestibular dark cells with the amiloride analog ethyl-isopropyl-amiloride (EIPA) and similar effects on Isc were observed with EIPA and the NHE blocker HOE694 when applied to the basolateral side of vestibular dark cell epithelium. The IC50 for these basolateral effects of EIPA, HOE694 and amiloride on Isc in vestibular dark cells were 2 x 10(-7) M, 8 x 10(-7) M and 4 x 10(-5) M. These observation suggest that vestibular dark cells and strial marginal cells contain NHE in their basolateral membrane, that K+ transport in strial marginal cells in pHi sensitive similar to K+ transport in vestibular dark cells and that NHE in vestibular dark cells consists of the subtype NHE-1.

Amiloride↗
PubMed2026

ECLIPSE: exploring the dark proteome of ESKAPE pathogens through the sequence similarity network of the Protein Universe Atlas.

MOTIVATION: The accelerating crisis of antimicrobial resistance among the critical so-called ESKAPE pathogens demands the urgent identification of novel molecular targets. However, a substantial fraction of ESKAPE proteomes remains functionally uncharacterized, with many genes annotated as encoding hypothetical proteins. These protein sequences often lack significant similarity to known protein families when conventional homology-based annotation methods are used and thus remain "dark". This limits our ability to explore their roles in pathogenicity, and it is thus crucial to bridge this substantial gap in pathogen biology by developing new strategies to illuminate these "dark" regions of the ESKAPE pan-proteome. RESULTS: We introduce ECLIPSE (ESKAPE Connectome Linkage and Inference for Proteome Sequence Exploration), a network-based computational framework that systematically identifies and prioritizes functionally dark protein families in ESKAPE pan-proteomes. ECLIPSE embeds target ESKAPE pathogen proteomes within the global sequence similarity network of the Protein Universe Atlas. It detects connected components composed entirely of unannotated proteins, called the "dark proteome." As a case study, we applied ECLIPSE to a pan-proteome of 3 460 657 protein sequences from 635 strains of Pseudomonas aeruginosa (PA). ECLIPSE identified 120 985 proteins (4%) residing in completely dark connected components. Furthermore, we have performed a taxonomic diversity analysis using normalized Shannon indices to characterize each dark component by its enrichment in ESKAPE pathogens. The analysis utilized the evenness (E) value (see Methods 2.1), which distinguishes Pseudomonas-specific (target-specific) from ESKAPE-enriched dark components. We then developed the Dark Proteome Prioritization Score (DPPS), a composite multidimensional scoring framework (see Methods 2.5). It ranks these dark components by biological relevance across four orthogonal axes: (i) functional darkness, (ii) P. aeruginosa proportion in the Atlas, (iii) AMR-clade taxonomic restriction, and (iv) conservation across the 635 P. aeruginosa strains. This framework outputs a robust four-tier scoring system; the prioritized Tier I components were validated by weight sensitivity analysis and remained stable across 500 Monte Carlo weight perturbations. Structural characterization of one of the top-ranked ESKAPE-enriched dark components revealed that it belongs to the beta-barrel fold DUF1302 (PF06980) family, for which no experimentally solved three-dimensional structure exists in the PDB. The genomic context analysis indicates that it is co-localized with a LuxR-type transcriptional regulator. Collectively, ECLIPSE identifies evolutionarily conserved, structurally defined, and functionally dark proteins enriched across ESKAPE pathogens; these dark proteins can further be utilized as alternative antimicrobial targets for experimental characterization. AVAILABILITY AND IMPLEMENTATION: The source code and dataset are available for free at: Github: https://github.com/surabhilata/ECLIPSE.git, Zenodo: DOI: 10.5281/zenodo.21064323.

Proteome↗
PubMed2000

Effects of light intensity and restraint on dark-pulse-induced circadian phase shifting during subjective night in Syrian hamsters.

Dark pulses presented on a background of constant light (LL) result in phase advances during midsubjective day and early subjective night, and phase delays during late subjective night, as shown in the dark-pulse phase response curve. In hamsters, the phase-shifting effects of dark pulses are thought to be mediated by increased activity, as previous studies have shown that restraining animals during dark pulses blocks the phase shifts observed in midsubjective day and late subjective night. This study focuses on dark-pulse-induced phase shifting during early subjective night, examining the influence of both LL intensity and restraint on the magnitude of these phase shifts. Syrian hamsters were maintained in LL of four different illumination levels (1, 10, 100, or 600 lux) and periodically presented with 6-h pulses (dark pulse alone, restraint alone, or dark pulse plus restraint) beginning at circadian time 11. Phase advances were observed in response to dark pulses alone, and the magnitude of these shifts was dependent on background illumination, with significantly larger advances seen under higher intensities. No relationship was found between the amount of activity displayed during dark pulses and phase shift magnitude. Six-hour periods of restraint resulted in phase delays, the magnitude of which was also dependent on background illumination. Restraining hamsters during dark pulses reduced the magnitude of phase advances, but the extent of this reduction could be predicted from the additive effects of the dark-pulse-alone and restraint-alone conditions. These results indicate that the phase-shifting effects of dark pulses during early subjective night are not mediated by behavioral activation and may instead reflect a mirror image of the phase-delaying effects of light pulses at this phase.

Animals↗
PubMed1988

Prostaglandins E1, E2, and D2 induce dark-adaptive retinomotor movements in teleost retinal cones and RPE.

In teleosts, retinomotor movements of photoreceptors and retinal pigment epithelium are regulated both by light and by an endogenous circadian rhythm. Light induces cones to contract, rods to elongate and RPE cells to disperse their pigment granules into their long apical projections; darkness induces opposite movements. When fish are maintained in prolonged constant darkness, appropriate movements nonetheless occur at subjective dusk and dawn. To explore the mechanisms of this light and circadian regulation, we have been investigating effects of several extracellular messengers known to be present in retina on retinomotor movements in the green sunfish (Lepomis cyanellus). Here we report that prostaglandin E1 (PGE1) can induce movements characteristic of dark onset (or night) in both cones and RPE in isolated light-adapted retinas in the light; ie, PGE1 induces cone elongation and RPE pigment granule aggregation. The extent of PGE1-induced cone and RPE movements were dose-dependent with maximal movement occurring at 250-500 nM; higher concentrations were not as effective. Incubations with PGE2 and PGD2 also induced dark-adaptive cone and RPE retinomotor movements, but PGF2 alpha did not. Further observations suggest that prostaglandins may play a role in mediating the induction of cone and RPE movements by dark onset: dark-induced movements were inhibited by pretreating light-adapted isolated retinas before dark culture with agents which inhibit endogenous prostaglandin synthesis. Both indomethicin (50 microM) and acetylsalicylic acid (50 microM), two inhibitors of the cyclooxygenase component of specific prostaglandin synthase, inhibited dark-induced cone elongation and pigment aggregation in cultured sunfish retinas. Another cyclooxygenase inhibitor, ibuprofen (50 microM) had no effect. Together the effectiveness of PGE1 inducing dark-adaptive movement and the inhibition of dark adaptive movement by cyclooxygenase inhibitors suggest that prostaglandins may play a role in vivo in mediating the induction of dark-adapted RPE and cone retinomotor movements by dark onset.

Alprostadil↗
PubMed2006

Plant-environment interactions: Accumulation of hypericin in dark glands of Hypericum perforatum.

BACKGROUND AND AIMS: Hypericum perforatum is a perennial herbaceous plant and an extract from this plant has a significant antidepressant effect when administered to humans. The plant is characterized by its secretory glands, also known as dark glands, which are mainly visible on leaves and flowers. The current study evaluates the influence of several environmental factors and developmental stages of the plant on the accumulation and synthesis of hypericin and pseudohypericin (Hy-G), the major bioactive constituents, in H. perforatum plants. METHODS: The appearance of dark glands on different parts of the plant, under several environmental conditions, was monitored by microscopy. Hy-G concentrations were quantified by high-performance liquid chromatography. KEY RESULTS: A significant presence of dark glands accompanying the highest concentrations of Hy-G was observed in the stamen tissues more than in any other organ of H. perforatum. A linear relationship between the number of dark glands and net photosynthetic rate of the leaf and Hy-G concentration in the leaf tissue was also established. A very high concentration of Hy-G was measured in the dark-gland tissues, but in the tissues without any dark glands it was almost absent. The presence of emodin, a precursor of Hy-G, at a high concentration in the dark-gland tissues, and its absence in the surrounding tissues was also observed, suggesting that the site of biosynthesis of Hy-G is in the dark-gland cells. A significantly low concentration of Hy-G (occasionally non-detectable) was measured in the xylem sap of the stem tissues. The dark-gland tissues collected from leaves, stems or flowers contained similar concentrations of Hy-G. CONCLUSIONS: The concentration of Hy-G in various organs of H. perforatum plants is dependent on the number of dark glands, their size or area, not on the location of the dark glands on the plant. The study provides the first experimental evidence that Hy-G is synthesized and accumulates in dark glands.

Anthracenes↗