Metabolism: the word to describe the totality of energy consuming, manipulative and storage chemical reactions by organisms. Second law of thermodynamics dictates that all processes increase amount of entropy in the universe. Thus, a highly ordered entity like a fish can only exist with a constant input of energy that allows it to remain ordered. Therefore, initial requirement of fish survival is to obtain sufficient energy to offset this universal randomization process by: maintaining ion gradients and renewing proteins (Chabot et al., 2016). “Respiration is nothing but a slow combustion of carbon and hydrogen, similar in all respects to that of a lamp or a lighted candle, and from this point of view, animals which breathe are really combustible substances burning and consuming themselves” (Lavoisier & Laplace, 1783). All animals must supply their cells with oxygen and rid their body of carbon dioxide. The physiological process by which an animal exchanges oxygen and carbon dioxide with its environment.Most fish have external gills that are ventilated by a unidirectional flow of water, by pumping or swimming. Fine sieve structure of gills very efficiently extracts O2 from water. Efficient O2 uptake is vital to fish because of its low water solubility. Solubility decreases with increased temperature & salinity! Also, metabolic rate (demand for O2 ) increases as temperature rises. ‘Branchia’ in greek = ‘gills’ In boney fish (Teleosts): Gills lie in a branchial cavity covered by the operculum: Usually two sets of four holobranchs. Each holobranch consists of two hemibranchs (‘half gill’): Anterior and posterior Hemibranchs consist of a row of long filaments (primary lamellae) with semilunar folds (secondary lamellae). Lamellae or filaments: Connective tissue scaffold (epithelial cells) framing a vascular network providing blood flow primarily used for gas and ion exchange.Fish employ the countercurrent system to extract O2 from the water. This system moves water flowing across the gills, in an opposite direction to the blood flow creating the maximum efficiency of gas exchange. Blood flow through lamellae is from posterior to anterior (back to front). Water flow over lamellae is from anterior to posterior (front to back). Counter-current allows for diffusion from high O2 in water to low O2 in blood across entire length of lamella.Gas gland is location of action in wall of swim bladder (rete mirabile “wonderful net” and surrounding tissues) Need to pry O2 molecules from Hb molecules in gas gland Need to accumulate enough O2 (>pO2) in solution in blood plasma to generate a diffusion gradient from distal end of rete mirabile into lumen of swim bladder Change of pH in blood causes change in bond strength of Hb for O2 Bohr effect--decrease in affinity of Hb for O2 due to decreasing pH or increasing pCO2 affinity: strength of attraction of Hb for O2 Root effect--decrease in capacity of Hb for O2 due to decreasing pH or increasing pCO2

1. Presented By: ASIFA WALI
Ph.D Scholar (II SEM)
Div: FRM
CREDIT SEMINAR- I
Gas transport in Fishes- A combination
of convection and diffusion

2. Dr. Tasaduq Hussain Shah (Major Advisor)
Dr. Farooz Ahmad Bhat
Dr. M.H. Balkhi
Dr. Bilal Ahmad Bhat
Dr. Tariq Ahmad Bhat (Dean PG Nominee)

3. CONTENTS
 INTRODUCTION
 METABOLISM IN FISHES
 OXYGEN CONSUMPTION IN FISH
 NATURE OF RESPIRATION
 FISH RESPIRATION
 TELEOST GILL ANATOMY
 NORMAL GILL HISTOLOGY
 TELEOST GILL STRUCTURE
 SCANNING ELECTRON MICROGRAPHS
 PROCESSES REMOVING AVAILABLE OXYGEN FROM
WATER…..

4. •Water is 840 times more dense
and 60 times more viscous than
air.
•Oxygen:
–Air: 210 ml/L O2 at 21%
partial pressure
–Water: up 15 mg/L O2
–Sea water holds 18% less
O2 than freshwater
•Oxygen consumption in fish
–17 mg/kg/h @10°C
–100-500 mg/kg/h @ 30°C
•More than 40 genera of fishes
breath oxygen using other
methods than their gills.
INTRODUCTION

5. Energy demand to accomplish
respiration: approx. 50% of
total demand but can be up to
90%.
Blood volume: 2-4 mL / 100 g;
(nucleated RBC)
Gill surface area: 150–300
mm2 / g tissue
Systolic blood pressure: about
44 mm Hg
Gill irrigation: 5-20 L H2O /
kg BM / h
Opercular beat counts: 40-60
/ minute
Life in water

6. METABOLISM IN FISHES
 Metabolism: the word to describe the totality of
energy consuming, manipulative and storage
chemical reactions by organisms.
 Second law of thermodynamics dictates that all
processes increase amount of entropy in the universe.
Thus, a highly ordered entity like a fish can only
exist with a constant input of energy that allows it to
remain ordered.
Therefore, initial requirement of fish survival is to
obtain sufficient energy to offset this universal
randomization process by: maintaining ion gradients
and renewing proteins (Chabot et al., 2016).

7. Metabolism of trout vs turtle
Trout Turtle
Oxygen requirement ~ 5 ml / min
/ kg
~ 5 ml / min
/ kg
Ventilation volume 600 ml H2O /
min / kg
50 ml air /
min / kg
Routine costs for
ventilation
10 % 2 %
Evans and Clairborne, (2006)

8. Metabolic rate in fish
 Standard metabolic rate (SMR)
 Routine metabolic rate (RMR)
 Active metabolic rate (AMR)
 Metabolic scope (MS): MS = SMR – AMR

9. 0.00
0.20
0.40
0.60
0.80
12:00 16:00 20:00 0:00 4:00 8:00
Hour of day
Oxygen
consumption
(mg.g
-1
.h
-1
)
AMR
SMR
9
Radull (2002)

10. “Respiration is nothing but a slow combustion of
carbon and hydrogen, similar in all respects to that of
a lamp or a lighted candle, and from this point of view,
animals which breathe are really combustible
substances burning and consuming themselves”
(Lavoisier & Laplace, 1783).
All animals must supply their cells with oxygen and
rid their body of carbon dioxide.
The physiological process by which an animal
exchanges oxygen and carbon dioxide with its
environment.
The Nature of Respiration

11.  Convective process of ventilation
supplies O2 to the gas exchange
surface.
 Diffusive processes then govern
the movement of oxygen across the
gill into the blood.

12. Most fish have external gills that
are ventilated by a unidirectional
flow of water, by pumping or
swimming.
Fine sieve structure of gills very
efficiently extracts O2 from water.
 Efficient O2 uptake is vital to
fish because of its low water
solubility.
 Solubility decreases with
increased temperature &
salinity!
 Also, metabolic rate (demand
for O2 ) increases as
temperature rises.

13. Oxygen solubility determined by temperature
Temp (C) O2 con. at sat.
(mg/l) – Fresh
O2 con. at sat.
(mg/l) – Salt
0 10.3 8.0
10 8.0 6.3
20 6.5 5.3
30 5.6 4.6
In warm water...fish need to extract MORE O2 from LESS!

14. Numerous lamellae protrude from both
sides of each filament and are the
primary sites of gas exchange.
Fish Respiration
Fishes use gills to extract oxygen from water: Counter-
current flow aids exchange.
Amphibians exchange gases across their skin, and at
respiratory surfaces of paired lungs: Larvae have external
gills.

15. The gills consist of bony or stiffened arches
(cartilage) that anchor pairs of gill filaments.
Gills are the main site of gas
exchange in almost all fishes.

16. Normal Teleost gill form (anatomy)
• ‘Branchia’ in greek = ‘gills’
• In boney fish (Teleosts):
• Gills lie in a branchial cavity
covered by the operculum:
• Usually two sets of four
holobranchs.
• Each holobranch consists of
two hemibranchs (‘half gill’):
• Anterior and posterior
• Hemibranchs consist of a row
of long filaments (primary
lamellae) with semilunar folds
(secondary lamellae).
• Lamellae or filaments:
• Connective tissue scaffold
(epithelial cells) framing a
vascular network providing
blood flow primarily used for
gas and ion exchange.
Secondary lamellae
Primary lamellae
Retrieved from the World Wide Web
http://www.biology-resources.com/drawing-fish-gill-filaments.html

17. Normal gill histology
Retrieved from the World Wide Web : http://aquaticpath.umd.edu/fhm/resp.html
Parasagittal section of the buccal cavity through the gill arches
(Bouins, H&E, Bar = 440 µm = ~ 4x magnification).
Head end
Gill rakers
Four paired
holobranchs
tongue
HOLOBRANCHS

18. Teleost gill structure
Schematic Diagram of the teleost fish gill.
Adapted from Evans et al. (2005)
Holobranch
Hemibranch (anterior)
Gill filaments

19. Scanning electron micrographs of:
(a) Branchial arch and filaments of gill from the teleost
(b) Filament and secondary lamellae of gill from the teleost
(a)
(b)

20. Healthy gills 2 layers of epithelial cells
Irritated gills - hyperplasia (reduction in exchange
efficiency)

21. 1) Short diffusion
distance at gill
site
2) Large surface
area for
diffusion at gill
site
3) Counter current
exchange of
gases at gill site
4) Large volume of
water passes
over gills
How can fish remove 80 - 90% of O2 available from water?

22. 3. Expel water from gill cavity
a. squeeze mouth
b. squeeze gill cavities open
operculum
4. Reset for next cycle
1. Fill mouth cavity
a. open mouth
b. expand volume of mouth
c. expand volume of gill
chamber with operculum
closed
2. Fill gill cavity
a. close mouth
b. squeeze mouth cavity
c. expand gill cavity with
operculum closed

23. Oxygen Exchange in Fish
 Fish employ the
countercurrent
system to extract O2
from the water.
 This system moves
water flowing across
the gills, in an
opposite direction to
the blood flow
creating the
maximum efficiency
of gas exchange.

24. gill filaments
water is sucked
into
mouth
Water exits
through gill
slits
A ) A teleost fish with its gill cover removed. Water flows in through the mouth,
flows over the gills, then exits through gill slits. Each gill has bony gill arches to
which the gill filaments attach.
one gill arch
Function of the gills of a Teleost

25. gill arch respiratory surface
gill
filament
fold with a
capillary
bed inside
water
flow
direction of
blood flow
oxygen-poor blood
from deep in body
oxygenated blood
back toward body
B
B) Two gill arches with filaments C
C) Countercurrent flow of water and blood

26.  Blood flow through
lamellae is from
posterior to anterior
(back to front).
 Water flow over
lamellae is from
anterior to posterior
(front to back).
 Counter-current allows
for diffusion from high
O2 in water to low O2 in
blood across entire
length of lamella.
Countercurrent
Close-up!

27. When the blood and water flows in the same direction, the co-current
system, it will initially diffuses large amounts of oxygen but the
efficiency reduces when the fluids start to reach equilibrium.
In the counter-current system, equilibrium is never reached!
Result: Oxygen flow is always directed into the gills.
% O2 in water
% O2 in gills
% O2 in water
% O2 in gills

28. Let’s Do the Math...
4 gill arches on each side of body
2 rows of gill filaments on each
arch (demibranchs)
100’s filaments per demibranch
- closely spaced
1000’s lamellae per gill filament
gill area = 10 to 60 times that of body
surface area, depending on species!
HUGE potential to extract Oxygen from water!

29. Branchial vs. Ram Ventilation
Branchial
 Mouth
 Pharynx
 Operculum
 Branchiostegals (filaments, lamella)
Ram
 Uses same parts, but not the pumping
energy required. Sharks primarily. Once
swimming speed is achieved...no need to
actively vent buccal cavity. However, this
can only be used consistently by strong
swimmers (sharks, tuna).
Shortfin Mako
(Isurus oxyrinchus)
Ram Ventilator
Pump Ventilator
Shortspine Spurdog
(Squalus mitsukurii))

30. GILL POUCHES

31. Auxiliary Respiratory Structures
 Skin - diffusion of oxygen from water into dense
network of capillaries in skin
 Swim bladder - vascularized physostomous swim
bladders
 Lungs - modified swim bladder
 Mouth - vascularized region in roof of mouth
 Gut - vascularized stomach or intestinal wall

33. Rete mirabile of a Queensland Grouper, Epinephelus
lanceolatus. © Geoff McPherson
 Gas gland is location of
action in wall of swim
bladder (rete mirabile
“wonderful net” and
surrounding tissues)
 Need to pry O2
molecules from Hb
molecules in gas gland
 Need to accumulate
enough O2 (>pO2) in
solution in blood plasma
to generate a diffusion
gradient from distal end
of rete mirabile into
lumen of swim bladder
 How?
Inflate a swim bladder
Gas bladder of a porcupine fish.

34. PCO2 / PO2 Lactate
pH Lactate
pH
Lactate
pH
Lactate
pH
PCO2 / PO2
PCO2 / PO2
PCO2 / PO2
Pre-rete Artery
Post-rete Vein
Gas
gland
epithelium
Secretory bladder
Bladder caps
Post-rete Artery
Pre-rete Vein
Rete mirabile: Changes of gas content, pH and lactate in the
capillaries
Rete
mirabile
Three processes:
• Bohr / Root shift
• Salting out
• Counter-current diffusion
O2
CO2
Evans and Clairborne (2006)

35. The rete counter-current system
Example: gas bladder rete in eel
• Cross section: 5 mm2
• Volume: 21 mm3
• Surface area: 30 cm2
• Capillaries: 20.000 to 40.000
• Artery diameter: 9 - 10 µm
• Venous capillary: 11 – 13 µm
• Diffusion distance (capillaries): 1
µm
• Capillary length: 4 mm
• Holes in capillary: 20 – 80 nm
• Hole diaphragm: 5 nm
Eye
Gas bladder
Ostrander (2000)

36. Prying O² from Hb
 Change of pH in blood causes change in bond
strength of Hb for O2
 Bohr effect--decrease in affinity of Hb for O2 due
to decreasing pH or increasing pCO2 affinity:
strength of attraction of Hb for O2
 Root effect--decrease in capacity of Hb for O2 due
to decreasing pH or increasing pCO2 (extreme
Bohr effect) capacity: total quantity of O2 that Hb
can carry
 more active species tend to have greater Bohr &
Root effects

37. Fish have nucleated red blood
cells!
Some species have more than 1
type of Hb
Hb exists in 2 states, a tense (T)
state with low affinity to O2, and a
relaxed state (R) with high affinity.
A shift from T -> R increases O2
binding capacity
The four units cooperate to increase
O2-uptake
What changes the state from T to
R?
Heamoglobin (Hb)

38. Carbon dioxide transport – the chemistry of CO2 in
water
CO2 H2O H2CO3 HCO3
- H+
+ +
Carbon dioxide
Carbonic acid
Bicarbonate ion

39. Extracellular
Carbonic anhydrase speeds up carbon dioxide
dissociation in the cell
CO2 H2O H2CO3 HCO3
- H+
+ +
CO2 H2O H2CO3 HCO3
- H+
+ +
Carbonic anhydrase
Red blood cell
Slow reaction
Fast reaction

40. Carbonic anhydrase speeds up carbon dioxide
dissociation at the tissues
CO2 H2O H2CO3 HCO3
- H+
+ +
Carbonic anhydrase
Fast reaction
Drop in pH !
Hemoglobin changes state
and releases O2
O2 diffuse to plasma
and tissues
HCO3
- transport to plasma
in exchange for Cl-

41. Carbonic anhydrase speeds up carbon dioxide
dissociation at the gills
CO2 H2O
H2CO3 +
Carbonic anhydrase
Fast reaction
Release of CO2 increases pH
Hemoglobin changes state and is ready to pick up
O2 at the gills.
CO2 diffusion to plasma
and water
HCO3
- H+
+
HCO3
- transport to blood
cell in exchange for Cl-

42. dissociation is slow
HHbO2 + O2
O2
CO2 + HbO2
HbCO2 + O2
O2
“Chloride Shift”

43. Partial pressure of oxygen (PO2)
Percent
saturation
of
Hb
with
O
2
higher pH
(gills)
lower pH
(tissues)
Bohr effect
Exponential
increase
due to Hb
subunit
cooperation
Dissociation
curves parallel
Reduced affinity under acidic conditions
Ostrander (2000)

44. Partial pressure of oxygen (PO2)
Percent
saturation
of
Hb
with
O
2
Bohr effect – species differences
Reduced affinity under acidic conditions
Ostrander (2000)
Sessile species with high Hb-affinity to oxygen
Can cope with low-oxygen conditions
Active species with low Hb-affinity to oxygen
Requires high oxygen levels to survive

45. Partial pressure of oxygen (PO2)
Percent
saturation
of
Hb
with
O
2
higher pH
(gills)
much lower pH
(retina / swimbladder)
Root effect
Subunit
cooperation
Some subunits
fail to load O2
O2 Saturation is not reached
even if sufficient O2 available!
Decreased capacity under acidic conditions

46.  To summarize O2 transfer from environment
to cells involve a series of steps; convective
process of ventilation, diffusive processes and
gas transport properties of blood togather with
convective process of blood flow deliver O2 to
tissues.
 The same processes determine transfer of
CO2 in opposite direction, from tissue site of
production to ventilatory water.
 To enable respiratory gas transfer to be
matched to variable metabolic demands of
tissues imposed by physical and environmental
stresses, a variety of strategies are available to
our fishes.
Most of these control strategies are keyed to
the tissues requirement for O2 probably because
O2 is difficult to obtain in an aquatic
environment owing to low O2 capacitance of
water in comparison to that of air.

47. THANK YOU

48. Chabot, D., Steffensen, J. F. & Farrell, A. P. (2016). The determination of standard metabolic rate in
fishes. Journal of Fish Biology (in press, this issue).
Evans, D.H., Piermarini, P.M. and Choe, K.P. (2005). The Multifunctional Fish Gill: Dominant Site of
Gas Exchange, Osmoregulation, Acid-Base Regulation, and Excretion of Nitrogenous Waste.
Physiological Review: 85, 97–177.
Evans, HE, Clairborne, JB (2006). The Physiology of Fishes, Third edition. CRC, Taylor and Francis.
Marine Biology Series.
Lavoisier, A. L. & Laplace, P. S. (1783). Memoire sur la chaleur. In Oeuvres de Lavoisier1862–1893, Vol.
2, pp. 283–333. Paris: Imprimerie Imperiale.
Ostrander, G.K. (2000). The Laboratory Fish. Academic Press.
Radull, J (2002). Investigations on the use of metabolic rate measurements to assess the stress response
in juvenile spotted grunter, Pomadasys commersonnii (Haemulidae, Pisces). PhD thesis, Rhodes
University.
Root, R.W. (1931). The respiratory function of the blood of marine fishes. Biological Bulletin of the
Marine Biology Laboratory, Woods Hole, 61, 427-457.
West, J. B. (2013). The collaboration of Antoine and Marie-Anne Lavoisier and the first
measurementsof human oxygen consumption. American Journal of Physiology. Lung Cellularand
Molecular Physiology 305, L775–L785.
REFFERENCES