PRIMARY RESEARCH PAPER

A fully implantable multi-channel biotelemetry systemfor measurement of blood flow and temperature: a firstevaluation in the green sturgeon

A. Grans Æ M. Axelsson Æ K. Pitsillides ÆC. Olsson Æ J. Hojesjo Æ R. C. Kaufman ÆJ. J. Cech Jr.

Received: 26 March 2008 / Revised: 17 August 2008 / Accepted: 24 August 2008

� Springer Science+Business Media B.V. 2008

Abstract The objective of this study was to evaluate

a novel fully implantable radio-based blood flow

biotelemetry system which allows simultaneously

measurement of blood flow on two channels and

temperature on one channel, in fish. These are the first

recordings of blood flow from free-swimming fish,

showing that the system is capable of recording blood

flow in the ventral aorta (cardiac output) and celia-

comesenteric artery (gastrointestinal blood flow) in

green sturgeon Acipenser medirostris exposed to a

series of different stimuli for up to 7 days after

implantation. The results showed stable base line

recordings and blood flow was used to calculated heart

rate (fH) and stroke volume (Vs). It was possible to

reproduce the same type of responses as has previously

been reported during exposure to hypoxia, tempera-

ture, stress and feeding. The mass of our implant was

less than 2% of the body mass which is well within the

recommended sizes for surgically implanted telem-

etry transmitters and it fitted easily within the abdom-

inal cavity of the sturgeon. A fully implantable system

minimizes the risk of infection/expulsion and maxi-

mizes the likelihood that the studied fish will behave

naturally and be treated normally by surrounding

fish. The use of biotelemetry in basic comparative

physiology and applied animal ecology could help

scientists to collect information that has previously

been challenging to obtain and to open the possibility

for new types of physiological and ecophysiological

studies.

Keywords Biotelemetry � Blood flow �Temperature � Doppler flow � Chronic measurements

Introduction

Biotelemetry systems are rapidly becoming important

tools for in vivo studies in medical and comparative

physiology. The major benefits of biotelemetry record-

ing systems, compared to traditional methods where

the experimental animals are physically connected to

the acquisition equipment through wires, are reduced

confinement stress, reduced risk of infections and the

possibility of long-term measurements in semi-natural

or natural environments. Biotelemetry makes it pos-

sible to record, for example, the effects of social

interactions and also to correlate normal behaviours

with physiological variables, something which is

Handling editor: K. Martens

A. Grans (&) � M. Axelsson � C. Olsson � J. Hojesjo

Department of Zoology, University of Gothenburg,

P.O. Box 463, 405 30 Goteborg, Sweden

e-mail: albin.grans@zool.gu.se

K. Pitsillides

EndoSomatic Technologies LLC, Sacramento, CA, USA

R. C. Kaufman � J. J. Cech Jr.

Department of Wildlife, Fish and Conservation Biology,

University of California, Davis, Davis, CA, USA

123

Hydrobiologia

DOI 10.1007/s10750-008-9578-7

practically impossible with ‘‘hardwired’’, cage-con-

fined animals. Many previous interpretations of

physiological data may be heavily affected or even

incorrect due to the stress induced by surgery,

confinement or human handling. The fact that the

animals studied using biotelemetry are no longer

physically connected to the recording equipment and

have the freedom to move around in a more natural

way may thus lead to higher quality of the data

obtained.

Biotelemetry is not a new idea. The first successful

transmission of biological information from a living

animal was performed in 1869 (Marey, 1896).

Winters (1921) was probably the first who success-

fully used radiotelemetry when he introduced the use

of a radiolink for sailors to get emergency advice

from doctors onshore when accidents happened at

sea. More than 20 years later, Fuller & Gordon

(1948) were the first to describe the use of biotelem-

etry to measure physiological activities from

unrestrained animals. When the transistor was intro-

duced in 1952 it became possible to design smaller

and more efficient biotelemetry equipment and this

led to a rapid increase in the use of telemetric

devices.

Blood flow is a central variable in cardiovascular

research and by measuring the blood flow of several

separate blood vessels (e.g. cardiac output and

gastrointestinal blood flow) information is obtained

about both the distribution of blood between tissues

and heart rate. Over the last 30 years there have been

several attempts to design an implantable blood

flowmeter, although none has been aimed for use in

fish. The new system is developed with the aim to

solve two central problems in fish biology: to

quantitatively describe the processing of food and

to evaluate the effects of stress on physiological and

behavioural mechanisms. These problems can be

addressed by measuring changes in distribution

between systemic blood flow and gastrointestinal

blood flow together with heart rate and temperature.

Several interesting systems have been described

that use different flowmeter techniques, such as,

electromagnetic flowmeters (Fryer et al., 1975),

ultrasonic Doppler shift techniques (Cathignol et al.,

1976; Allen et al., 1978, 1979) and interferometric

ultrasonic techniques (Rader et al., 1975). However,

most system, have parts of the devices externally

mounted and are consequently limited for use in

larger animals (Franklin et al., 1964; Yonezawa

et al., 1989, 1992; Spelman et al., 1991; Kong et al.,

2007). Other problems with these devices are the high

power consumption (Fryer et al., 1975; Cathignol

et al., 1976; Allen et al., 1978, 1979) and the bulk

flow probes (Rader et al., 1975).

Recently the first paper describing a multi-channel

biotelemetry system capable of recording both blood

pressure and blood flow was presented (Axelsson

et al., 2007). This multi-channel flow and pressure

biotelemetry system was designed for larger animals

and has so far been used and tested in pigs and

alligators (Axelsson et al., 2007). The system has

capacity to simultaneously measure blood flow at

four channels and blood pressure at three channels

together with ECG and temperature. This resulted in

a fairly large system (around 130 g, 60 cc volume)

(Axelsson et al., 2007), precluding application for

fish. However, based on this larger multi-channel

system a smaller device has been developed and the

present study includes the first evaluation of this

system. In the smaller system two of the blood flow

channels are retained together with one channel

allowing measurement of temperature. Due to the

difficulty to accurately measure blood pressure when

the external pressure fluctuates, e.g. when the freely

swimming fish changes depths, pressure was not

recorded in the smaller version. Our dual-channel

system consists of two parts: a component that is

fully implanted in the fish and a base station. Similar

to the larger system, the dual-channel system uses a

bidirectional communication protocol between the

base station and the implant. This allows not only

data to be received and processed from the implant,

but also commands to be sent to the implant.

Two important aspects during the development of

either system were the requirements for a fairly long

transmission range and a relatively long battery

lifetime. Long range transmission means that the

base station/computer and implant can communicate

with each other which allows the fish under study

more freedom compared to hardwired animals that

are usually restricted to relatively small experimental

chambers.

The evaluation study was conducted on green

sturgeon, Acipenser medirostris (Ayres), which is

native to North America. The choice of species

allowed us to compare our new data with older

studies using traditional methods, looking at the same

Hydrobiologia

123

cardiovascular variables in sturgeon, conducted in the

same laboratory (Crocker et al., 2000). The sturgeon

were available at different size ranges facilitating

determination of the appropriate size of animals for

the implant.

The aim of this study was to test the newly

developed dual blood-flow and temperature biote-

lemetry system in free-swimming fish. Data obtained

from telemetrically instrumented animals exposed to

temperature challenges, hypoxia, chasing and feeding

in the laboratory were compared with data obtained

from earlier studies using traditional techniques.

Material and methods

Animals used

The green sturgeon (n = 3) used in this study were

3 years old and had the masses 3.3 (sturgeon #1), 3.6

(sturgeon #2) and 4.2 (sturgeon #3) kg. They were

descendants of wild-caught Klamath River stur-

geon that were artificially spawned in 2004 (Van

Eenennaam et al., 2001). The eggs were incubated at

the University of California, Davis, at the Center for

Aquatic Biology and Aquaculture (CABA) and the

juveniles were kept in air-equilibrated water at tem-

peratures similar to the Klamath River (11–15�C)

during late spring. The fish were fed commercial

Silvercup trout pellets at 3–5% body weight ration per

day based on a feeding table for white sturgeon

Acipenser transmontanus (Richardson) that also has

been successfully used for green sturgeon (Kaufman

et al., 2006). At 31 days post-hatching, the fish were

placed in round 284-l fibreglass holding tanks and kept

there for their first 2 years, and then transferred to

12,800-l tanks until needed for experiments. The tanks

received a continuous flow of air-equilibrated, 19�C

well water.

Short description of the newly developed system

A simplified block diagram of the system is presented

in Fig. 1. This fully implantable biotelemetry system

consists of two channels for Doppler-based flow

measurements and one channel for temperature

measurement. The Doppler flowmeter is designed to

reduce power consumption through micro-power

analog and radio-frequency integrated circuits, as

well as advanced power management techniques. For

a further description see Axelsson et al. (2007). The

power reducing design results in a system which can

run on a single AA-size lithium battery with all three

channels operating continuously for approximately

14 days. Table 1 summarizes the technical specifica-

tions of the biotelemetry system. The implantable

flowmeter is also capable of timed-acquisition oper-

ation where the average power consumption is further

reduced. The timed-acquisition operation controls the

turning on and off of the implant in repeated, timed

intervals, thus substantially extending battery life.

The implant stores in internal memory its calibra-

tions for the flow and temperature channels. These

calibrations are easily accessed from the base station

user interface by sending a calibration command to the

implant. The implant responds with a 3-step calibra-

tion signal for the two flow channels and a 2-step

calibration signal for the temperature. The bidirec-

tional transceiver range is approximately 2–6 m

depending on enclosure, water depth and surrounding

radio frequency (RF) interference.

Implant description

The implant is controlled by a low-power microcon-

troller that generates all power control and timing

signals to the Doppler flowmeter and temperature

subsystems. The microcontroller also encodes and

formats the data for transmission through the RF link,

and processes any received commands. The Doppler

Flow 1Flow 2Temp

Trigger

RS232

RFLink Digital to

AnalogConverters

Controller

Battery

Flow 1Flow 2

Temp TempSensor

RFLinkController

DopplerFlowmeter

Module

A

B

Fig. 1 Block diagram of (A) base station and (B) implant

Hydrobiologia

123

flowmeter module operates at 20 MHz. Each channel

has independent adjustment controls for range-gate

positioning, signal direction and channel on/off.

Temperature is measured using an internal, digital

sensor.

Base station description

There are three basic functions that are performed in

the base station decoder/controller unit:

1. Reception and decoding of data from implant.

2. Transmission of user-selected commands and

configuration parameters to the implant.

3. Conversion of decoded implant data to analog

signals (using the digital-to-analog section). The

control of the implant/base station is mediated

via the computer using a custom-made program

and a serial communication protocol between the

computer and the base station.

The analog output from the base station was

connected to a commercial data acquisition system

(ML865 PowerLab 4/25T Data acquisition System,

AD Instruments Pty Ltd, Castle Hill, Australia)

running the software Chart 5 for Windows on a

PC-computer (Dell Latitude 820). The sampling rate

was set to 100 Hz for all channels.

Preoperative care

The sturgeon were anaesthetized by placing them in a

40-l well water tank containing 0.2 g l-1 3-amino-

benzoic acid ethyl ester (MS-222), 10 g l-1 NaCl and

buffered to pH 7.0 with 4.2 g l-1 NaHCO3. The

sturgeon were kept in the anaesthetics until ventila-

tory movements ceased and then weighed and

measured before being transferred to an operating

table covered with wet sponges. The fish were

positioned with the ventral side facing up and the

eyes shielded from the bright light using wet paper

towels. Anaesthesia was maintained by pumping

oxygenated, buffered MS-222 (0.075 g l-1) over

the gills during the surgery. A sterile surgical drape

(1051 Incise Drape, 3 M, USA) was used to cover

the fish and to keep the surgical surface sterile and

sterile gloves (Sensi-Touch Latex Powdered Sterile

Surgeons Glove, Ansell Healthcare Products Inc.,

Canada) were used throughout the surgery. Before

the surgery, the fish were injected with 2.5 mg/kg

[1 M] Enrofloxacin (Baytrilo�, Bayer, USA) antibi-

otics and all surgical instruments, the implant and

associated electrical leads were sterilized by the use

of a cold sterilant (Cidex, Johnson & Johnson

Company, USA). The surgical precautions taken

have been stressed in several reviews on fish surgery

(Butcher & Wildgoose, 2001; Fontenot & Neiffer,

2004) and were decided together with veterinary staff

at the University of California (UC), Davis. All

animal experiments were performed in accordance

with national and local ethical guidelines (EH&S

protocol: 07-12677).

Gastrointestinal blood flow and cardiac output

Green sturgeon have a single large celiacomesenteric

artery (CoMe) that supplies the entire gastrointestinal

tract with blood. To expose the celiacomesenteric

artery, a 5-cm-long midventral incision was made

posterior to the pectoral girdle and the liver was

carefully retracted. Gastrointestinal blood flow

(qCoMe) was recorded using silicon cuff transducer

probe with two integral canals for insertion of sutures

Table 1 A summary of the specifications for the system

Dimensions and battery life

Implant dimensions 60 9 30 9 10 mm

Battery pack dimensions 50 mm, Ø15 mm

Total weight 45 g

Battery life 340 h continuously ON

Doppler flowmeter

Number of channels 2

Ultrasound frequency 20 MHz

Pulse repetition frequency 64 kHz

Minimum range gate adjustments 1 mm

Maximum range gate adjustments 6 mm

Doppler frequency shift measured [18 kHz

Blood velocity range [100 cm/s

Transducer diameter 0.7 mm

Temperature

Number of channels 1

Temperature range 0–50�C

Resolution 0.0625�C

Radio frequency link section

Frequency 433 MHz

Power out 0 dBm

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123

(ES-2.5, Iowa Doppler products, Iowa City, USA)

that were placed around the CoMe. The probe cuffs

were closed using the integrated silk sutures.

In order to measure cardiac output (Q), a 2-cm

ventral incision was made posterior to the gill

juncture and carefully, without disrupting the peri-

cardium or damaging any vessels, the dermal and

sub-dermal musculature and connective tissue were

separated using blunt dissection tools to expose the

ventral aorta. A silicon, cuff-type Doppler blood flow

transducer with two integral canals for insertion of

sutures (ES-4.0, Iowa Doppler products, Iowa City,

USA) was placed around the ventral aorta. To

internalize the lead from the ventral aortic flow

probe, the probe was tunnelled under the skin from

the ventral incision to the midventral incision. The

probe was then placed around the ventral aorta and

closed using the integrated silk suture. The wires

from the probes were anchored with a single stitch of

3/0 silk suture in the sub-dermal muscle tissue. The

implant and battery were then carefully placed in the

abdominal cavity and the retracted organs restored to

their places of origin. The two incisions were closed

using sterile 3/0 nylon monofilament suture. The

locations of the probes and the implant are illustrated

in Fig. 2.

The surgical procedure including anaesthesia and

awakening took between 60 and 90 min.

Postoperative care

Post-operatively the sturgeon were given a subcuta-

neous injection of 0.1 mg/kg Torbugesic (butorphanol,

Fort Dodge, Iowa, USA) for post-operative pain relief,

in accordance with veterinarian instructions. The fish

were moved to 1.3-m-diameter fibreglass tanks

(750 l), where they could move freely, for recovery.

The tank received a continuous flow of air-equili-

brated, 19�C well water. Fish were allowed at least

24 h to recover from surgery before experimental

exposures started.

Experimental protocol

Cardiac output (Q), gastrointestinal blood flow (qCoMe)

and temperature (T) were continuously recorded

during the tests. Heart rate (fH) was calculated from

the phasic cardiac output signal using the cyclic

Q

qCoMe

Implant

battery

Fig. 2 Arrangement of the

implant, battery and the two

Doppler flow probes that

recorded cardiac output (Q)

and gastrointestinal blood

flow (qCoMe) inside the

sturgeon. The implant and

battery were positioned in

the abdominal cavity and

the flow probes measuring

Q and qCoMe were placed on

the ventral and

celiacomesenteric arteries,

respectively

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123

measurement feature in the Chart 5 software, while

stroke volume (Vs) was calculated from Q and fH.

During the experiments, all sturgeon were exposed

to three separate stimuli (hypoxia, increased water

temperature and chasing) once. Before each stimulus

all variables were recorded for at least 1 h to ensure

stable baseline values. The order of the stimuli was

randomized and the fishes were allowed at least 24 h

recovery between exposures. Precautions were made

to minimize long-term stress for the sturgeon in the

experiment. Similarly, the chasing exposure was

designed to give short-term effects. Chasing is a

comparably mild stimulus compared with other

previously used methods such as netting and water

reduction (Lankford et al., 2005), minimizing the

recovery time needed, and therefore reducing the risk

of carry-over effects between trials.

Lankford et al. (2005) showed in a study on

juvenile green sturgeon that plasma levels of two

indicators of stress, cortisol and glucose, were never

elevated for more than 180 min after a single stressor.

However, chronic stress (2–3 stressors randomly each

day) caused elevated levels of plasma glucose. More

than three stressors a day were not possible since it

causes increased mortality rate (Cech, unpublished).

In our study we wanted to measure blood flow

responses caused by several stimuli, with minimal

long-term effects to minimize the carry-over effects

between the different days and exposures. It has been

shown that green sturgeon stressed during nights

show higher levels of plasma cortisol and glucose

than do sturgeon stressed during the day (Lankford

et al., 2003). Consequently, all exposures were con-

ducted between 06:00 and 18:00 to minimize the

recovery times. It has also been shown that temper-

ature may affect recovery after stressful events.

Green sturgeon kept at 19�C recover from a 1-min

air emersion stressor faster (based on plasma cortisol

levels) compared to animals exposed to the same

stressor but acclimated to a lower temperature of

11�C (Lankford et al., 2003). Hence, the fish were

kept at 19�C, except during the temperature

challenge.

Temperature exposure

Heated (30�C), air-equilibrated well water was

pumped into the holding tank at a flow of 1.8 l min-1,

leading to a temperature increase of approximately

1.25�C every 30 min. The temperature was allowed to

increase from 19 to ca. 26�C after which the heated

water was turned off and the water temperature

returned to pre-test values.

Hypoxia exposure

Nitrogen gas (N2) was introduced via gas diffuser

stones into the holding tank and partial oxygen

pressure (PO2) was measured every 20 min using an

oxygen electrode (E101 Cameron oxygen electrode,

Analytical Sensors, Inc, Sugar Land, TX, USA). The

flow of N2 was increased stepwise from 20 to

40 ml min-1 to decrease PO2 from the initial value

of 120 mm Hg down below 60 mm Hg before the N2

flow was turned off and the water in the tank slowly

returned to normoxia.

Chasing

The sturgeon were manually chased with a net for

15 min. Chasing as a general stressor has previously

been used as for shortnose sturgeon, Acipenser

brevirostrum (Lesueur) (Beyea et al., 2005) and a

5-min chasing session also has been used previously as

a stressor for green sturgeon (Lankford et al., 2005).

Feeding

After the three stimuli described above one of the

animals was lightly anaesthetized, placed on its back

and fed by gavage. The meal consisted of commercial

Silvercup trout pellets equivalent to 1% of the body

weight. This is a common method in laboratory based

feeding studies and makes it possible to control for

meal size and exact time of feeding. The postprandial

effects on Q and qCoMe were then followed contin-

uously for approximately 33 h.

Results

Validation of technique

Green sturgeon were ideal to use for validating this

new system. Surgery was fast and minimal and they

seemed unaffected by the implants. Signals were

strong and clear despite movements in the tanks.

During the chasing all the animals were very active,

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but no deterioration of the signal quality could be

seen, indicating a robust signal transmission.

Figure 3 shows a longer (60-min) section of

continuous recording of Q and qCoMe, in sturgeon

#1, 24 h after implantation. From the two traces of

raw data, a 30-s section is enlarged to verify the

quality of the signals. The left part of the lower

panels shows the unfiltered signal, while the right

shows the signal after filtering using a Triangular

Bartlett window (window width: 25 points). During

the exposures, adjustments in the setting were done

manually which resulted in some variations in the

duration of each step in the treatments. Hence, only

the response of one sturgeon to each stimulus is

presented in the figures (Figs. 4–7).

Temperature

Figure 4 shows the original recording of sturgeon #3

during the temperature challenge. Only a moderate

increase in Q, Vs and fH occurred when the temper-

ature were raised from 19 to 24�C. In contrast, qCoMe

increased by more than 150% when the temperature

increased above around 23�C. The response differed

among the three tested fishes with two animals

showing a decreased qCoMe (\50%) when the tem-

perature was increased above 25�C. Between 19 and

24�C, all three fish showed similar responses and

mean values (change in response at 24�C in percent

of response at 19�C) are presented in Table 2. The

fish body temperature was obtained from the implant

0

40

80

120

0

20

40

Blo

odF

low

(cm

s-1

)

Raw data Same trace Smoothed*

* Triangular (Bartlett), window width: 25 points 10 s

10 min

0

0

20

100

Blo

odF

low

(cm

s-1

)

A

BQ

qCoMe

Q

qCoMe

Fig. 3 Verification of signal quality from the two flow probes.

(A) Raw data for cardiac output (Q) and gastrointestinal blood

flow (qCoMe) during 60 min continuous recording in a resting

sturgeon under control conditions. (B) A section of the trace in

A corresponding to 30 s with raw data for cardiac output (Q)

and gastrointestinal blood flow (qCoMe) shown on the left and

the same trace after filtering using a Triangular Bartlett,

window (window width: 25 points) on the right

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123

and water temperature measured with an additional

thermometer showed no lag time between the

increase in water temperature and fish temperature.

Hypoxia

Figure 5 shows the data from sturgeon #2 during the

hypoxia challenge. In this animal, Q, Vs and fHincreased when PO2 got below 80 mm Hg. The

opposite was seen in qCoMe where the flow decreased

95% compared with the initial value. Also during the

hypoxia trial the responses differed among individual

sturgeon. In sturgeon #3, bradycardia appeared when

PO2 was dropped to approx. 50 mm Hg. Mean values

for all animals comparing the responses at PO2 of 120

and 60 mmHg are presented in Table 2.

Chasing

Figure 6 shows the response from sturgeon #2 to the

15 min net chasing. Q, Vs and fH showed an increase

when the chasing started. Q and Vs increased ca. 20%,

but again the largest effect was seen in qCoMe, which

decreased 95% compared with the initial value. A

comparison of the responses prior and during chasing

is presented as mean values for all three animals in

Table 2.

Tem

p(°

C)

20 min0

17

19

21

23

25

27

0

100

200

300%

VsqCoMe

Q

T

f H(m

in-1

)

0

20

40

60 fH

Fig. 4 Trace from a temperature (T) challenge obtained from

sturgeon #3. The temperature was increased from 19 to 26�C

during a period of approximately 2 h. When the temperature

increased moderate increases occurred in cardiac output (Q),

stroke volume (VS) and heart frequency (fH), while the

gastrointestinal blood flow (qCoMe) increased by more than

150%

15 min

0

50

100

150

200

0

20

40

60

50

75

100

125

0

fH

PO2

f H(m

in-1

)%

PO

2(m

mH

g)

VsqCoMe

Q

Fig. 5 Trace from a hypoxia challenge obtained from sturgeon

#2. Cardiac output (Q), stroke volume (VS) and heart frequency

(fH) increased and the gastrointestinal blood flow (qCoMe)

decreased when partial oxygen pressure (PO2) declined below

80 mm Hg

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Feeding

Figure 7 shows the feeding trial from sturgeon #2.

This animal showed an increase in Q during the first

3 h after feeding, followed by a return to a value

close to what was observed before feeding. The blood

flow to the gastrointestinal tract initially decreased

followed by an increase with a peak around 12 h

postprandially. At this time the gastrointestinal blood

flow had increased by ca. 75% compared with values

prior to feeding and by approx. 125% compared with

values 2 h after feeding.

Discussion

To the best of our knowledge this is the first study to

record blood flow from fish using a fully implantable,

radio-based biotelemetry implant. It shows that the

newly developed system is capable of recording

blood flow in the ventral aorta and celiacomesenteric

artery in free-swimming sturgeon exposed to several

different stimuli.

The electronic part of this dual channel system is

the same as used by Axelsson et al. (2007). In that

study, extensive tests were conducted in order to

evaluate the correlation between the signal from the

implant and the signal obtained from a Triton

Instruments model 100 Doppler flowmeter which is

a traditional benchtop equipment. A strong correla-

tion (r2 = 0.978) was seen indicating that signal

quality was not deteriorated. Furthermore, by visual

inspection the quality of the signals from the two flow

probes in the present study appeared similar to that

from a previous study, using ultrasonic blood flow

probes in white sturgeon (Crocker et al., 2000). This

demonstrates that the telemetric system is capable of

reproducing signals equally to standard benchtop

equipment.

The external stimuli used in this study (increased

temperature, hypoxia and chasing) are all challenges

which green sturgeon may face in their natural

habitats. Increasing temperature and hypoxia are

natural phenomena with which all fish living in

temperate habitats need to cope.

Heart frequency was calculated from Q, and levels

were comparable to values reported for both white

sturgeon (Crocker et al., 2000) and Siberian sturgeon,

Acipenser baerii (Brandt) (Maxime et al., 1995).

Both hypoxia and chasing elicited a drastic decrease

in qCoMe; this was also seen during the temperature

exposures when the temperatures were at the upper

end of the tested range. The responses seen in the

present study were both quantitatively and qualita-

tively comparable to the results obtained in an earlier

study of the white sturgeon (Crocker et al., 2000)

Chasing5 min

0

50

100

150

0

10

20

30

40

50

60 fH

f H(m

in-1

)%

VsqCoMe

Q

Fig. 6 Trace from a chasing challenge obtained from sturgeon

#2. The challenge consisted of a 15-min net chasing. Cardiac

output (Q), stroke volume (VS) and heart frequency (fH)

increased and gastrointestinal blood flow (qCoMe) decreased

drastically

5 h

Feeding 1 % of body mass

Blo

odflo

w(%

)

0

100

200

50

150

qCoMe

Q

Fig. 7 Trace from forced feeding of sturgeon #2. Cardiac

output (Q) remained elevated 3 h after feeding. The gastroin-

testinal blood (qCoMe) flow initially decreased followed by an

increase which peaked around 12 h after feeding

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123

with the difference that the animals were swimming

freely without any physical connection to any

recording equipment in the present study.

A reduction in blood flow to the gastrointestinal

tract during exercise (induced swimming or chasing)

has been described in various species: for example,

Chinook salmon, Oncorhynchus tshawytscha

(Walbaum) (Thorarensen et al., 1993; Thorarensen

& Farrell, 2006) and rainbow trout, O. mykiss

(Walbaum), (Randall & Daxboeck, 1982). A decrease

in qCoMe during hypoxia has also been shown in

several species including European sea bass, Dicen-

trarchus labrax (L.), (Axelsson et al., 2002) and

Atlantic cod, Gadus morhua (L.) (Axelsson & Frit-

sche, 1991). All these studies support the hypothesis

that blood is distributed away from the stomach when

there is a need to optimize oxygen delivery, either due

to a reduction in available environmental oxygen

(hypoxia) or during exercise when there is an

increased demand from swimming muscles that needs

to be met by the cardiovascular system.

During hypoxia, a bradycardia was observed in

sturgeon #3 at the lowest partial oxygen pressure.

Hypoxic bradycardia is a well-documented response

in fish, including the closely related Adriatic sturgeon

where bradycardia developed at deeper levels of

hypoxia (Agnisola et al., 1996). Hypoxic bradycardia

is discussed in several reviews but is still not

completely understood (Randall, 1982; Farrell, 2007).

In the feeding test the sudden decrease in qCoMe

and increase in Q after feeding was most likely due to

stress caused by handling at the time of feeding.

Hyperemia in the gastrointestinal canal induced by

feeding is expected when activities such as gut

motility, intracellular biochemical activity and mem-

brane transport all increase after feeding (McCue,

2006). This has been described in several fish species

including sea raven, Hemitripterus americanus

(Gmelin), (Axelsson et al., 1989), red Irish lord,

Hemilepidotus hemilepidotus (Tilesius), (Axelsson

et al., 1989), Atlantic cod (Axelsson & Fritsche,

1991), European sea bass (Axelsson et al., 2002) and

Chinook salmon (Thorarensen et al., 1993). Due to

differences in temperature, amount of food given,

species-specific differences and effects of instrumen-

tation between the studies it is difficult to make direct

comparisons. However, in all cases feeding induces

an increase in blood flow to the gastrointestinal canal.

In some species this increase is provided by an

increase in cardiac output, while in other species

there is a redistribution of blood flow, leaving cardiac

output unchanged. The dynamics of the changes are

highly dependent on temperature and effects of

instrumentation. Axelsson et al. (2002) showed in

their study on the effects of feeding and hypoxia in

the European sea bass that instrumentation signifi-

cantly increased the gastric evacuation time. This is

an area where implantable telemetric systems will be

useful with the associated, increased possibility for a

successful recovery.

Few studies have successfully managed to link

natural behaviour with changes in blood flow due to

the difficulty in getting instrumented or in other ways

restrained fishes to behave naturally. The use of

biotelemetry in basic comparative physiology and

applied animal ecology could help scientists to

collect information that has previously been chal-

lenging to obtain and to open the possibility for new

types of physiological and ecophysiological studies

(Cooke et al., 2004).

Pros and cons with blood flow biotelemetry?

A perfect biotelemetry device should be very small

and fully implantable so that it will not affect the

bearer. The size of the biotelemetry implants, relative

to the animals to be studied, has been and is still one of

the largest problems when developing the systems.

Table 2 Response in cardiac output (Q), gastrointestinal blood flow (qCoMe), stroke volume (VS) and heart frequency (fH) to

increased temperature, hypoxia or chasing in green sturgeon (Acipenser medirostris)

Q qCoMe VS fH

Temperature 23.44 ± 16.48 45.28 ± 22.86 -5.78 ± 11.26 29.87 ± 2.99

Hypoxia -4.63 ± 16.80 -63.19 ± 15.10 3.57 ± 8.89 8.45 ± 9.79

Chasing 10.54 ± (5.37) -75.73 ± 11.80 7.22 ± 4.54 3.20 ± 1.67

Values are mean ± standard deviation from all three sturgeons, presented as percent change compared with the baseline response

(19�C, 120 mm Hg and before chasing, respectively) at peak response (24�C, 60 mm Hg and after chasing, respectively)

Hydrobiologia

123

The general recommendation is that the implant

should not exceed 2 to 5% of the body mass of the

animal, although the appropriate forms and mass

should be evaluated for each species because size and

morphology vary enormously (Jepsen et al., 2002).

The mass of our implant was \2% of the body mass

and the implant fitted easily within the abdominal

cavity of the sturgeon. Our system is also the first fully

implantable system capable of measuring blood flow

in fish. A fully implantable system minimizes the risk

of infection and expulsion (Schulz, 2003) and also

maximizes the likelihood that the studied fish will

behave naturally and be treated normally by sur-

rounding fish (Connors et al., 2002). Even the

presence of a small, protruding antenna has been

shown to elicit aggressive attacks from other individ-

uals of Atlantic salmon, Salmo salar (L.), smolts

(Connors et al., 2002).

Long-term effects of implants are often evaluated

using dummy implants (Connors et al., 2002; Jepsen

et al., 2002; Schulz, 2003). Since the primary goal of

the current study was to look at the functionality of

this newly developed blood flow telemetric system in

fish, this was not considered necessary at this stage.

However, the long-term effects of implantation will

be studied in more detail in future studies.

Until now the commercially available biotelemetry

equipment used in fish has been transmitters designed

to measure fH and electromyograms (EMG). EMG

telemetry has been an essential tool in studies that

have helped us, e.g. to understand better the migra-

tion and spawning ecology of sockeye salmon

Oncorhynchus nerka (Walbaum) (Hinch & Rand,

1998; Hinch & Bratty, 2000; Hinch et al., 2002;

Healey et al., 2003).

Estimating metabolic rate from blood flow

Metabolic rate (VO2) is the basis of life and the single

most important variable in a living system. Metabolic

rate is a function of cardiac output and arterio-venous

oxygen content. Several attempts have been made to

obtain information about VO2 in free-swimming fish,

based on fH. This was first described by Priede and

Tytler (1977) and has thereafter frequently been

used in other studies although with mixed results

(Armstrong et al., 1989; Lucas & Armstrong, 1991;

Lucas et al., 1991; Lucas, 1994; Armstrong, 1998;

Lefrancois et al., 1998). Because stroke volume and

heart rate can change independently, the use of heart

rate as an indicator of cardiac output has to be

considered carefully. For a somewhat reliable estimate

of VO2 from fH, one must make a laboratory calibra-

tion to obtain fH/VO2 correlation curves which can be

used in a field situation. It has been shown that the use

of fH as an indicator of metabolic rate is limited to

stable environments and that the VO2 values obtained

in the laboratory from confined animals may compare

poorly with those from fishes in their natural environ-

ments (Claireaux et al., 1995; Thorarensen et al.,

1996; Webber et al., 1998). In their study on Atlantic

cod, Webber et al. (1998) showed that, if the fH/VO2

correlation-curves are based on the recovery period,

VO2 can be overestimated by as much as 100%. For a

more detailed discussion describing the limitations of

fH as a predictor of VO2, see reviews by Thorarensen

et al. (1996) and Butler et al. (2004).

Q is a more reliable variable for prediction of VO2

because both fH and VS are considered. Therefore, a

biotelemetric system that simultaneous measures both

blood flow and heart rate would minimize the

problems associated with fH-based VO2 estimates

(Thorarensen et al., 1996; Webber et al., 1998;

Altimiras & Larsen, 2000). However, problems also

can arise from estimating VO2 from Q (Brodeur

et al., 2001), and it is important to stress that

correlations among Q, fH and VO2 may differ

between species and environments (Thorarensen

et al., 1996; Armstrong, 1998; Lefrancois et al.,

1998; Webber et al., 1998; Altimiras & Larsen,

2000; Brodeur et al., 2001).

Heart rate, obtained with biotelemetry devices, has

also been used to estimate variables such as activity

and digestion (Armstrong, 1986), although this

method also has been criticized for being biased. In

a study conducted on European sea bass, it was

shown that following stress or during digestion, fish

metabolic turn-over rates temporarily reached values

close to active VO2 (Lefrancois et al., 1998). Taking

into account the described problems with laboratory-

derived correlation curves based on fH values, with

the expected tachycardia response to digestion,

exercise and stress, the difficulty to separate the

relevant factors in a uncontrolled situation is obvious.

By using two separate blood flows it will be possible

not only to measure changes in both fH and VS but

also to measure regional blood flows (e.g. the blood

allocated to the gastrointestinal tract).

Hydrobiologia

123

The effect of welfare on data quality

Another important and interesting aspect of using

biotelemetric equipment instead of hardwired animals

is the possibilities for prolonged recovery after

surgery and reduced stress from handling. The time

needed for a fish to recover from surgery depends on

species, individuals and environment (Fontenot &

Neiffer, 2004). Most studies only allow a recovery

period of 12–48 h after surgery before onset of the

experiment. However, several studies have shown

elevated fH levels for much longer and have con-

cluded that 48 h of recovery after surgery may be too

short (Webber et al., 1998; Altimiras & Larsen, 2000;

Campbell et al., 2005). Webber et al. (1998) describe

a 2–7-day period for physiological recovery in

Atlantic cod. Altimiras & Larsen (2000) showed

how the metabolic scope decreased when rainbow

trout recovered for up till 2 days after surgery.

Therefore, short recovery periods might lead to

misinterpretations and false conclusions. One of the

main reasons for the commonly used short recovery

time is the high risk of infection after an extensive

surgery (Fontenot & Neiffer, 2004). Much of histor-

ical data might be biased by the poor health of

the studied fishes (Altimiras & Larsen, 2000;

Huntingford et al., 2006; Johansen et al., 2006). In

a study using bioelectric potential recordings to

measure fH, rainbow trout were handled and then left

to recover for at least 3 days. The basal heart rates of

these trout were much lower than those presented in

previous studies using traditional methods involving

surgical procedures (Altimiras & Larsen, 2000).

Long-term measurements using biotelemetry would

help to verify or reject proposed basal values for

physiological parameters. The focus in this study was

to test the system and to evaluate the quality of the

data obtained by comparing the results with data from

studies using traditional bench-test equipment on

hardwired animals. The aspects of recovery time on

the recorded variables and the possible differences

in response to the various challenges should be

examined in a future study.

Future applications for the system

It is clear that the biotelemetry system can record

cardiac output and gastrointestinal blood flow from

free-swimming sturgeon (at least in a laboratory

environment) and that the responses obtained with

this system compare well with results from earlier

studies. In this study the aim was primarily to

evaluate the functionality of the system and there was

no possibility to look at the long-term effects in the

measured variables if the animals were allowed full

recovery, something which is impossible with tradi-

tional recording methods. With the increased use of

fish models in most biological disciplines (Powers,

1989), the need for animal welfare guidelines asso-

ciated with animal models in research has emerged

(Harms, 2005; Johansen et al., 2006). The telemetry

system can also help the fish culture industry. With

increasing commercial values of farmed fish, it is of

both ethical and economical interests to keep healthy,

unstressed animals (Conte, 2004; Ashley, 2007). If

fish are stressed and the responses, including loss of

appetite, impaired growth and muscle wasting are

prolonged, the consequences for the fish farmers can

be devastating (Huntingford et al., 2006). Developing

a management protocol to reduce stress and its

consequences is a shared goal in aquaculture (Conte,

2004). Today there are several ways to monitor

cultured fish’s welfare, looking at both environmental

variables, such as levels of oxygen and carbon

dioxide, and physiological variables. Most physio-

logical variables are monitored by visual cues such as

changes in coloration or behaviours of the fish

(Huntingford et al., 2006; Ashley, 2007). Although

trials have been conducted using biotelemetry and

include measurements of muscular activity, gill

ventilation rate and heart rate, questions concerning

how culture conditions affect the fish’s health remain

unanswered (Baras & Lagardere, 1995). Each year

the aquaculture industry loses massive amounts of

money on problems related to stress and the farmers

are requesting new ways of monitoring the health of

the fish (Conte, 2004; Ashley, 2007).

With this novel biotelemetry system, a possibility

to conduct long-term monitoring in cultured fish

emerges. By monitoring both total cardiac output and

gastrointestinal blood flow together with temperature

within an aquaculture setting, it would be possible to

devise systems and operating procedures to minimize

stressful situations, optimizing fish growth rates and

production.

Another area where biotelemetric studies may

increase our understanding is the effects of climate

changes on fish populations. It has been shown that

Hydrobiologia

123

populations of some marine fishes are moving

northwards as ocean temperatures increase (Roessig

et al., 2004; Perry et al., 2005). Migration towards

colder northern water is possible in many marine

environments but may not be a solution for freshwa-

ter species, living in lakes and river systems.

Although Ficke et al. (2007) documented stream

fishes, moving to higher elevations, some populations

will have to either adapt or adjust to the changes

instead. If this is not possible, their scope for activity

and reproduction will be reduced, potentially jeopar-

dizing their survival. The seasonal differences and

shifting environments caused by global warming

alters both the physiology and behaviour of many

freshwater species (review by Farrell, 1997). The

effects of climate-induced changes are only possible

to study partially in the laboratory; it is important to

get information from animals in their natural envi-

ronment and biotelemetric systems are the only

possible way to obtain physiological data in this

situation. Because temperature changes directly

affect the metabolism of fish (and all other ectother-

mic animals) and because blood flow is a much better

estimate of metabolism compared with heart rate, the

biotelemetry system described in this paper could

be a useful tool in assessing effects of temperature

changes in fish, caused by either natural seasonal

variations or anthropogenic effects (Randall &

Brauner, 1991).

Conclusion

The first evaluation of this novel biotelemetric system

shows that it is capable of replicating results obtained

using traditional benchtop equipment. We show that

blood flow measurements are possible to conduct

with animals swimming in their ‘‘home’’ tanks

instead of in confined cages. To evaluate the advan-

tages of this system as compared with traditional

methods, further studies are needed.

Acknowledgements This research was supported by

Goteborg University Research Platform on Integrative

Physiology (GRIP), the Swedish Science Research Council

Grants B-DR 09856-304 and 621-2002-3869 (M. Axelsson),

the Helge Ax:son Jonsson foundation and the Knut & Alice

Wallenberg foundation. We gratefully acknowledge the

assistance of W. Wesley Dowd, Christa M. Woodley and the

rest of the staff at Department of Wildlife, Fish and

Conservation Biology, University of California, for adjusting

their time plans to make this study possible. K. Pitsillides owns

EndoSomatic Technologies, LLC, which developed the

biotelemetry system described in this manuscript and for

more information on the biotelemetry system visit

www.endosomatic.com.

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