Functional hypoxia : induced by high activity.
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Transcript of Functional hypoxia : induced by high activity.
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Functional hypoxia: induced by high activity.
Environmental hypoxia: surrounding PO2 decreases.
,
Hypoxia
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Oxidative phosphorylation contribution to ATP production
Facultative Anaerobe
Oxygen Regulator
ATP turnover
Anoxia
Hypoxia
Unlimiting O2
^
Glycolytic contribution
Oxidative phosphorylation contribution to ATP production
Metabolic depression is a key adaptation in intertidal marine invertebrates and it allows long-term hypoxia/anoxia tolerance
Oxygen regulators undergo a Pasteur effect (top), but facultative anaerobes demonstrate a “reverse Pasteur effect” (bottom).
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Oxidation toCO2 and H2O
Glucose
Pyruvate Lactate
NAD+NADH + H+
NAD+
NADH + H+
G3PDH
Glucose utilization under AEROBIC and ANAEROBIC conditions.
NADH+ + H+
NAD+
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Fermentations Energy yield H+ yieldGlucose Lactate: 2ATP/Glu 1ATP/H+
Glucose Opine: 2ATP/Glu 1ATP/H+
Glucose Succinate: 4ATP/Glu 2ATP/H+
Glucose Propionate: 6ATP/Glu 3ATP/H+
Aspartate Succinate: 1ATP/AspAspartate Propionate: 2ATP/Asp
Marine invertebrates have multiple anaerobic pathways
There are also H+ consuming metabolic processes:
Adenylate Deaminase reaction: AMP + H2O IMP + NH3 (forms NH4+)
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Cellular Energetics Under Hypoxia/Anoxia
4 reaction systems are linked (3 supply, 1 demand):
Aerobic metabolism: Glucose + ADP + Pi ATPCreatine kinase rxn: PCr + ADP + H+ ATP + CrGlycolysis: Glucose + ADP + Pi ATP + Lactate + H+
ATPase: ATP ADP + Pi
Net effect of energy challenge:↓PCr, ↑Cr, ↓Glucose, ↑Lactate, ↓pH, ↑ADP, ↑Pi
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Phospho-creatine
Pi
Nuclear magnetic resonance can be used to assess cellular energetics non-invasively
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31P-NMR analysis of muscle energetics in vivo in goldfish (A) (ethanol producers) and tilapia (B) (lactate producers).
Note the decrease in PCr and increase in Pi during hypoxia. Also note the Pi peak has shifted to the left during hypoxia (decreased pH).
The ATP levels remain fairly constant but G of ATP hydrolysis declines.
Intracellular energetics in fish during hypoxia
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20
30
40
50
60
High High-Mid
Mid Low
Intertidal Position
Buf
feri
ng C
apac
ity
(um
ol/p
H/m
l) 30 mM acidload
60 mM acidload
0.1
0.2
0.3
0.4
0.5
0.6
High High-Mid
Mid Low
Intertidal Position
dH+/d
t (um
ol/m
l/min
)
30 mM acidload
60 mM acidload
Buffering capacity and the capacity for proton pumping is correlated with intertidal position in closely related intertidal whelks.
Intracellular buffering of H+: Previously shown that buffering capacity was correlated with energy demand in fish (functional hypoxia) (Castellini and Somero, 1981). Is habitat related to the ability to regulate pH?
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Portner et al. 2000. J. Exp. Biol.
Extracellular pH decreases reduce the cost of membrane transport processes associated with pH regulation. Inhibitors: Ouabain (Na/K ATPase), DMA (amiloride, Na/H antiport), DIDS (anion exchange, such as Na+ dependent Cl-/HCO3
- exchange), Bafilomycin (V-ATPase H+ pump inhibitor).
Cellular pH regulation can be costly
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Mackerel (fast, active, open water)
Scup (inshore still waters)
Toadfish (sluggish, inshore)
Innes and Taylor, 1986 Stewart, 1991
Gas exchange surfaces and hemoglobin binding affinity are dependent on environment and behavior
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Wood, 1991Brix et al. 1989
Euphausiid P50 is higher at lower T. Goldfish move to lower T when PO2 drops, which lowers metabolic rate and may help unload O2 at tissues.
Other environmental factors can alter hemoglobin P50
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Fish respond by invoking anaerobic metabolism and increasing the ventilation rate
Respiratory and metabolic response to hypoxia in the epaulette shark, which lives on reef platforms that become hypoxic at low tide. The pattern is typical of most vertebrates, lactate increases and VO2 decrease once a critical PO2 is reached. Ventilation rate is increased to compensate for the reduced oxygen (Routley et al. 2002).
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How is oxygen sensed by cells?
Hypoxia-induced gene expression via HIF. HIF1 is constitutively expressed. A heme-based receptor detects O2 levels and leads to an oxygen-dependent modification of HIF1. This modification allows HIF1 to be ubiquinated, which targets it for destruction. When HIF1 is not modified, it can dimerize with ARNT to form a heterodimer transcription factor that can bind enhancer sites on DNA.
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mRNA from tissue/cell
cDNA (fluorescently label)
Hybridize to an array
Determine genes that are up-regulated (green), down-regulated (red), or unchanged (yellow).
Control TreatmentMicroarray analyses of Response to Hypoxia
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Hypoxia-induced gene expression in the hypoxia-tolerant goby Gillichthys mirabilis examined using cDNA microarrays. PO2 was approximately 10% of normoxia, which is below PO2 that induces a decrease in respiration. Gene expression was then measured in a control, and at times after hypoxia exposure (Gracey, Troll and Somero, 2001).
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ATP metabolism
locomotion
translation
iron metabolism
anti-growth/proliferation
amino acid metabolism
cryptic role
Northern Blots
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A Special Case: The Diving Response in Marine Mammals
From Hochachka and Somero (2002)
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Field studies of the Weddell seal showing increase in hematocrit during diving. Also, in the field, the lactate washout peak was often small or absent during short dives, and large during long dives. This led to the concept of the aerobic dive limit (ADL).
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Heme
His F8
Hydrophobicpocket
In vivo elephant seal NMR spectra during normal breathing (A), sleep apnea (B) and post-apnea normal breatning (C). The apnea duration was 1-10 min and the deoxy-Mb peak emerged after 3 min. The stability of the [Pi] (1), [PCr] (2), and [ATP] (3-5) suggest that Mb unloading supports oxidative phosphorylation during apnea (Ponganis et al. 2002).
1H-spectra 31P-spectraMyoglobin Function
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Top: [Mb] (bars) and maximum dive duration (circles) for species of cetaceans and pinnipeds.
Bottom: Body mass relationship to maximum dive duration.
(Noren and Williams, 2000).
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How do marine mammals exceed the aerobic dive limit without becoming anaerobic?
Top: Dive depth vs. duration in marine mammals showing gliding time (red) and swimming time (black).
Bottom: Recovery oxygen costs (repaying the oxygen debt) in the Weddell seal for a gliding dive is less than for a swimming dive. A gliding dive extends the aerobic dive time by 38%.
Williams et al. (2000)
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Glycolytic flux in RBCs from marine and terrestrial mammals during a 2 h incubation under high hydrostatic pressure (Castellini et al. 2001).
Are marine mammals sensitive to pressure?