Respiratory Physiology & Neurobiology 181 (2012) 249–258
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Respiratory Physiology & Neurobiology
journal homepage: www.elsevier.com/locate/resphysiol
Developmental changes in cold tolerance and ability to autoresuscitate from
hypothermic respiratory arrest are not linked in rats and hamsters
Andrea E. Corcoran a,∗ , Denis V. Andrade b , Lieneke H. Marshall c , William K. Milsom c
a
b
c
Department of Physiology and Neurobiology, Dartmouth Medical School, Lebanon, NH, USA
Departamento de Zoologia, Universidade Estadual Paulista, Rio Claro, SP, Brazil
Department of Zoology, University of British Columbia, Vancouver, BC, Canada
a r t i c l e
i n f o
Article history:
Accepted 8 March 2012
Keywords:
Respiratory control
Postnatal development
Cold tolerance
Autoresuscitation
Rat
Hamster
a b s t r a c t
In adult mammals, severe hypothermia leads to respiratory and cardiac arrest, followed by death.
Neonatal rats and hamsters can survive much lower body temperatures and, upon artificial rewarming,
spontaneously recover from respiratory arrest (autoresuscitate), typically suffering no long-term effects.
To determine developmental and species differences in cold tolerance (defined here as the temperature
of respiratory arrest) and its relation to the ability to autoresuscitate, we cooled neonatal and juvenile
Sprague-Dawley rats and Syrian hamsters until respiration ceased, followed by rewarming. Ventilation
and heartbeat were continuously monitored. In rats, cold tolerance did not change throughout development, however the ability to autoresuscitate from hypothermic respiratory arrest did (lost between
postnatal days, P, 14 and 20), suggesting that the mechanisms for maintaining breathing at low temperatures was retained throughout development while those initiating breathing on rewarming were altered.
Hamsters, however, showed increased cold tolerance until P26–28 and were able to autoresuscitate into
adulthood (provided the heart kept beating throughout respiratory arrest). Also, hamsters were more
cold tolerant than rats. We saw no evidence of gasping to initiate breathing following respiratory arrest,
contributing to the hypothesis that hypothermic respiratory arrest does not lead to anoxia.
© 2012 Elsevier B.V. All rights reserved.
1. Introduction
Hypothermia, a large unregulated decrease in body temperature
(TB ), is a lethal condition for most mammals. Once a critically low
TB is reached, respiration ceases and is followed by cardiac arrest
and ultimately death if the animal is not rewarmed. Early work by
Adolph (1951) showed that most species of adult mammals, including rats, could not withstand body temperatures below 15–20 ◦ C,
although in some species (such as hamsters) adults were able to
survive a TB as low as 5 ◦ C. He also found that neonates had an
increased cold tolerance compared to their adult counterparts and
both newborn rats and hamsters could tolerate temperatures as
low as 1 ◦ C (Adolph, 1951). Clearly, while adults lose cold tolerance,
this is more profound in some species than others. Adolph (1948a,b)
also found that neonates cooled until both respiration and cardiac arrest occurred, could spontaneously resume both rhythms on
rewarming (autoresuscitation). Adults, however, could not; they
required ventilatory assistance to resuscitate.
∗ Corresponding author at: Department of Physiology and Neurobiology, Dartmouth Medical School, 1 Medical Center Drive, Lebanon, NH 03756-0001, USA.
Tel.: +1 603 650 6387; fax: +1 603 650 6387.
E-mail address: [email protected] (A.E. Corcoran).
1569-9048/$ – see front matter © 2012 Elsevier B.V. All rights reserved.
doi:10.1016/j.resp.2012.03.006
Several researchers have now documented the developmental decline in cold tolerance (defined as lower lethal temperature
in such studies) (Adolph, 1948a,b, 1951; Adolph et al., 1961;
Fairfield, 1948; Hill, 2000) and found that, in small mammals, it
occurs predominantly between 10 and 20 days after birth. The
time frame during which the ability to spontaneously recover from
hypothermic respiratory arrest is lost, however, has not yet been
documented. It has been shown that during lethal hypothermia,
respiratory arrest occurs first and that the temperature threshold
for this event is different from that for cardiac arrest (Hill, 2000).
Thus, while the loss of the ability to autoresuscitate is undoubtedly
related to the loss of cold tolerance, the relationship between these
two is not clear. Thus in the present study we sought to determine the relationship between cold tolerance and the ability to
autoresuscitate from hypothermic respiratory arrest as well as the
developmental time frame during which changes in these variables
occur in both rats and hamsters.
Many developmental changes in cardio-respiratory processes
have now been observed in rats around postnatal days 10–12
(P10–12) including changes in chemosensitivity (Davis et al., 2006;
Putnam et al., 2005; Wang and Richerson, 1999), responses to acute
hypoxia (Liu et al., 2006), and neurochemical changes in brainstem
nuclei involved in respiratory control (Wong-Riley and Liu, 2005,
2008). The second week of life appears to be a critical period for
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Table 1
Mean values of select variables during cooling and warming in rats.
Mass (g)
fR at 35 ◦ C
fH at 35 ◦ C
Rate of cooling (◦ C/min)
Rate of re-warming (◦ C/min)
TB at respiratory arrest (◦ C)
fH at respiratory arrest (beats/min)
TB when breathing resumes (◦ C)
fH when breathing resumes (beats/min)
Survival (%)
P0–6a (27)
P1 (4)
10.8 ±
93 ±
349 ±
0.45
0.71
10.7 ±
41.2 ±
13.3 ±
27.4 ±
100
28.0 ±
103 ±
311 ±
0.37 ±
0.44 ±
9.4 ±
28.4 ±
13.5 ±
27.6 ±
75
0.6
4
7
0.24
15.0
0.4
2.3
P16 (4)
1.5
13
29
0.03*
0.0 (3)
2.0
11.5
2.8 (3)
10.8 (3)
34.3 ±
161 ±
373 ±
0.29 ±
0.41 (1)
9.9 ±
23.9 ±
16.8 (1)
29.0 ±
25
P18–20 (4)
±
±
±
±
1.3
92
21
0.04
38.1
98
388
0.21
1.9
21
54
0.03
1.5
9.5
8.3 ± 1.7
26.1 ± 7.3
8.8 (1)
0
Numbers in parenthesis = n values.
a
Data for P0–6 rat pups from Tattersall and Milsom (2003).
*
Significantly different from P16 and P18–20.
Table 2
Mean values of select variables during cooling and warming in hamsters.
Mass (g)
fR at 35 ◦ C
fH at 35 ◦ C
Rate of cooling (◦ C/min)
Rate of re-warming (◦ C/min)
TB at respiratory arrest (◦ C)
fH at respiratory arrest (beats/min)
TB when breathing resumes (◦ C)
fH when breathing resumes (beats/min)
Survival (%)
P2–3 (9)
P5–6 (9)
P8–9 (8)
P15–20 (3)
P26–28 (6)
P32–34 (7)
4.5 ±
53 ±
405 ±
0.49 ±
0.66 ±
8.5 ±
38.5 ±
20.3 ±
78 ±
100
7.8 ±
65 ±
420 ±
0.46 ±
0.60 ±
4.9 ±
39.2 ±
13.1 ±
23 ±
100
10.4 ±
58 ±
388 ±
0.47 ±
0.57 ±
3.8 ±
25.7 ±
10.8 ±
24 ±
100
16.9 ±
109 ±
474 ±
0.25 ±
0.37 ±
3.5 ±
13.5 ±
9.3 ±
17 ±
100
45.9 ±
206 ±
515 ±
0.22 ±
0.42 ±
3.6 ±
18.6 ±
11.0 ±
18 ±
100
52.5 ±
148 ±
259 ±
0.65 ±
0.93 ±
8.9 ±
17.0 ±
10.0 ±
4±
71.4b
0.2
2
15
0.02
0.06
0.07
5.7
2.1
15
0.3
6
10
0.02
0.04
0.4
5.0
2.1
3
0.5
6
12
0.02
0.04
0.3
4.4
1.0
4
3.1
28
29
0.04
0.03
0.4
0.6
0.6
2
2.6
22
3
0.02
0.03
0.4
7.2
0.6
4
2.2
35a
12a
0.05
0.11 (5)
0.6
1.2
0.6 (5)
1 (5)
Adults (8)
127.9 ±
195 ±
337 ±
0.35 ±
0.53 ±
6.0 ±
19.3 ±
7.7 ±
16 ±
62.5b
6.3
20a
24a
0.03
0.02 (5)
0.7
3.6
1.7 (5)
5 (5)
Numbers in parenthesis = n values.
a
Data recorded at 30 ◦ C rather than 35 ◦ C.
b
Animals not surviving also went into cardiac arrest.
respiratory development (Wong-Riley and Liu, 2008). Accordingly,
we hypothesized that the time course over which the switch from
the neonatal to the adult phenotype for hypothermic respiratory
arrest would occur in the rat would correspond with this time
course. Furthermore, since it has been shown that hamsters are
more cold tolerant than rats (Adolph, 1951), and that in vitro
brainstem–spinal cord preparations from hamsters exhibit respiratory arrest and autoresuscitation at lower temperatures than
Fig. 1. Breathing frequency during transitional cooling (filled circles) and subsequent re-warming (open circles) in neonatal rats aged P0–6, P14, P16, and P18–20. Data are
presented as absolute means ± SEM. Data for rats aged P0–6 were obtained from Tattersall and Milsom (2003).
A.E. Corcoran et al. / Respiratory Physiology & Neurobiology 181 (2012) 249–258
251
Fig. 2. Breathing frequency during transitional cooling (filled circles) and subsequent re-warming (open circles) in (A) neonatal and juvenile hamsters aged P2–3, P5–6, P8–9,
P15–20, P26–28, P32–34, and (B) adult hamsters. Data are presented as absolute means ± SEM.
those of rats (Fong et al., 2009; Zimmer and Milsom, 2004), we also
hypothesized that this temporal window would occur later in the
hamster.
2. Methods
2.1. Animals
Experiments were conducted on neonatal and juvenile SpragueDawley rats (Rattus norvegicus) and Syrian hamsters (Mesocricetus
auratus). Animals were divided into groups based on postnatal age
(P, days after birth). Rats were grouped into P14, P16, and P18–20.
We have previously published data for P0–6 rats (Tattersall and
Milsom, 2003). Hamsters were grouped into P2–3, P5–6, P8–9,
P15–20, P26–28, P32–34, and P71–73 (adults). Rats were obtained
from the University of British Columbia (UBC) Animal Care Centre the morning of an experiment. Pregnant Syrian hamsters were
purchased from Charles River Rodent Laboratories (Calgary, AB,
Canada) and allowed to give birth naturally. Hamster pups were
randomly separated from their mother immediately prior to an
experiment. All experiments were completed under the guidelines
of the Canadian Council for Animal Care and with prior approval
from UBC Animal Care Committee.
2.2. Animal instrumentation
Animals were anesthetized with 1–2% halothane. Those >P10
possessed fur and were shaved on the ventral, dorsal and lateral
body surfaces. This minimized insulation and therefore thermoregulatory defense during cooling, insuring a similar rate of cooling
of animals at different developmental stages. Disk electrodes for
measuring the electrocardiogram (ECG) and respiratory impedance
were inserted beneath the skin on both the upper left chest and
lower right abdomen, and the incisions sealed with tissue glue
(Periacryl). A copper–constantan thermocouple wire attached to
a digital display monitor (Physitemp model BAT-12) was inserted
into the rectum to record body temperature. All wires leading from
electrodes were secured to the animal’s back using medical tape or a
piece of latex glued to the skin. Once the animal was instrumented it
was placed inside a water-jacketed, temperature-controlled chamber (set to 35.5 ◦ C).
2.3. Measurements of ventilation
Breathing was assessed differently depending on animal size
and activity (these variations were largely due to the extensive age
range used). The following three techniques were used to monitor breathing: (1) head-out body plethysmography, (2) whole-body
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Fig. 3. Heart rate during transitional cooling (filled circles) and subsequent re-warming (open circles) in neonatal rats aged P0–6, P14, P16, and P18–20. Data are presented
as absolute means ± SEM. Data for rats aged P0–6 were obtained from Tattersall and Milsom (2003).
plethysmography, and (3) impedance plethysmography. Note that
due to technical barriers, it was difficult to accurately measure tidal
volume (VT ), therefore VT was recorded in only a subset of animals.
For head-out plethysmography, a flexible piece of latex with
a small aperture was fitted around the animal’s neck, thus
providing a seal separating the body and head into different
temperature-regulated compartments (Mortola, 1984). For whole
body plethysmography, the entire animal was placed inside a single sealed chamber. Air (warmed to a parallel temperature to
that inside the compartment) was passed through the chamber
at a constant rate that ensured adequate ventilation. Air pressure
changes associated with chest wall movements within the chamber were detected by a pneumotachograph attached to a Validyne
differential pressure transducer. Calibration of the differential pressure detected by the pneumotachograph was achieved by injecting
known volumes of air into the chamber and measuring the voltage
change that resulted (Mortola, 1984). The third method of obtaining breathing traces measured the impedance (resistance) across
the chest wall of the animal using the electrocardiogram electrodes.
With this technique a small amount of alternating current, too small
to be detected by the animal, was passed through the chest wall.
The resistance to the current increased during inspiration, as the
lungs were filled, and decreased during expiration, as the lungs
were emptied. This change in resistance was measured with an
impedance converter (UFI, Model 2991). The electrode and thermocouple wires exited the plexiglass chamber through a port sealed
with plasticine, permitting the animal to move freely inside. Heart
rate and ventilation signals were amplified (through a Gould AC
and DC amplifier, respectively), filtered and recorded continuously
on a WindaqPro computer data acquisition system.
2.4. Experimental protocol
Once instrumented, animals were placed in the chamber for
∼30 min prior to cooling. Euthermic values were recorded for
10 min after which cooling was initialized by setting the temperature of the water-jacket to 5 ◦ C. Body temperature fell gradually
(0.2–0.7 ◦ C/min). After approximately 60–180 min, depending on
the animal’s size and age, breathing arrested. Respiration was considered to have arrested once the animal did not breathe for 10
consecutive minutes. This was usually when body temperature
had fallen to between 4 and 8 ◦ C. The TB of the animal was kept
0–2 ◦ C below the temperature at which arrest occurred for 10 min.
Subsequently, the animal was slowly re-warmed (0.35–1 ◦ C/min)
by increasing ambient temperature until its body temperature
reached euthermic values. At the end of an experiment, animals
were euthanized with an overdose of halothane followed by decapitation.
2.5. Data and statistical analyses
Breathing frequency (fR ) and heart rate (fH ) were determined
for each degree TB during cooling and re-warming and the mean
for each age group of each species was calculated. The mean TB at
which respiratory arrest occurred, mean TB of autoresuscitation,
and also the rates of survival were determined for each age group
of each species. Tidal volume (VT ) and inspiratory time (TI ) were
determined for (a) first breath after respiratory arrest (upon rewarming) (b) average of 5 breaths at the same TB during cooling
as the first breath upon re-warming and (c) average of 5 breaths at
35 ◦ C (Fig. 8). We included (b) as a comparison to minimize temperature as a confounding variable potentially affecting VT and TI .
Paired t-tests were used to examine significant differences between
mean TB of arrest and mean TB of autoresuscitation. A one-way
ANOVA was used to test for effects of age on the TB of arrest, and
a one-way ANOVA on ranks was used to test for effects of age on
the TB of autoresuscitation. To test for significant differences in air
flow rates of breaths, a one-way repeated measures ANOVA was
performed on the slope of VT /TI .
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253
Fig. 4. Heart rate during transitional cooling (filled circles) and subsequent re-warming (open circles) in (A) neonatal and juvenile hamsters aged P2–3, P5–6, P8–9, P15–20,
P26–28, P32–34, and (B) adult hamsters. Data are presented as absolute means ± SEM.
3. Results
3.1. Cooling to the minimum TB
Rats and hamsters were cooled to a temperature at which
breathing arrested taking between 70 and 165 min. Average
rates of cooling for both species ranged from 0.21 ± 0.03 to
0.65 ± 0.05 ◦ C/min (Tables 1 and 2). In rats, the rate of cooling was
slower in the two older groups reflecting their increased size. In
hamsters, however, although body weights also increased with age,
there did not appear to be consistent changes in rates of cooling or
warming that would reflect larger animals taking longer to cool or
re-warm. This was also despite, that for the first 15 ◦ C of cooling,
both rats and hamsters older than P15 exhibited rigorous shivering
in response to the cold.
3.2. Breathing frequency and heart rate during cooling
During initial cooling between TB 35 ◦ C and 20–25 ◦ C, with the
exception of P2–3 hamsters and P0–6 rats, fR remained relatively
constant (Figs. 1 and 2). Below ∼20–25 ◦ C, fR decreased in a roughly
linear fashion in all age groups of both species until respiratory
arrest occurred (Figs. 1 and 2). In all hamsters and most rats, the
relationship between fH and TB was linear from the onset of cooling until a minimum of ∼10 beats/min was reached (Figs. 3 and 4).
As a result, from Figs. 5 and 6 it can be seen that in the youngest
animals, both breathing frequency and heart rate fall in concert but
as the animals age, breathing frequency is maintained while heart
rate continues to fall during cooling at temperatures above roughly
20 ◦ C.
3.3. Respiratory arrest and autoresuscitation
Rats arrested breathing at similar temperatures across all ages
examined (average TB of arrest for rats P14–20 was 9.1 ± 0.94 ◦ C)
(Fig. 7). In hamsters, however, age was a factor in temperature of respiratory arrest: P2–3 and P32–34 stopped breathing at
significantly higher temperatures than other age groups. The temperature of arrest dropped slowly until P26–28 and then increased
again. Note that hamsters ceased breathing at lower temperatures
than rats of comparable ages (P = 0.016; t-test). The mean TB of
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Fig. 5. The relationship between breathing frequency and heart rate during transitional cooling (filled circles) and subsequent re-warming (open circles) in neonatal rats
aged P0–6, P14, P16, and P18–20. Data are presented as absolute means ± SEM. Data for rats aged P0–6 were obtained from Tattersall and Milsom (2003). Arrows indicate
values during cooling at 20 ◦ C.
autoresuscitation in hamsters was consistently higher than the TB
of arrest for any given age.
While 100% of neonatal P0–6 rats autoresuscitated in the previous study (Tattersall and Milsom, 2003), we observed that of
P14 rats, only 75% autoresuscitated from hypothermia-induced
respiratory arrest whereas only 25% of P16, and none of P18–20
rats autoresuscitated. The developmental change in the ability to
recover from hypothermia-induced respiratory arrest was species
dependent as all hamsters younger than P32–34 autoresuscitated.
At P32–34, autoresuscitation was limited to 71.4% and only 62.5%
of adult hamsters recovered completely from respiratory arrest.
If, however, P26–28 hamsters were left at the minimum TB for
30 min of no breathing instead of the usual 10 min, they failed to
recover (n = 3, data not shown). Note, however, that cardiac arrest
also occurred in all hamsters that failed to autoresuscitate. If their
hearts continued to beat at very low temperatures, they were destined to recover, despite the length of respiratory arrest (average
of 25.5 min in surviving adults).
Similarly, age effects on the TB at which animals took their first
breath on re-warming (TB autoresuscitation) from the minimum
TB was species dependent (Fig. 7). Irrespective of age, rats destined to autoresuscitate spontaneously resumed breathing during
re-warming at a similar temperature mean for the 4 animals that
recovered was 13.7 ± 1.1 ◦ C, which is comparable to the value of
13.3 ± 0.38 ◦ C reported for neonatal rats aged P0–6 by Tattersall and
Milsom (2003). This was not the case for the hamsters where the
TB at which breathing resumed decreased with age until P15–20
and remained at this level in all older age groups (Fig. 7). Note
that TB of autoresuscitation was generally higher than the TB
at which animals underwent respiratory arrest in rats (with the
exception of P0–6) and younger hamsters. In the older hamsters
(>P32) the TB at which breathing arrested and resumed were not
different.
3.4. Breathing frequency and heart rate during rewarming
The first breaths upon rewarming in both rats and hamsters
(P14–20 and P26–28, respectively) were significantly longer in
duration than those in normothermy (35 ◦ C) and those at the same
temperature on cooling. Apparent increases in VT failed to meet statistical significance (Fig. 8). The airflow rates during the first breaths
on rewarming (VT % of control/TI ) (Fig. 8) in juvenile rats were similar to the rates of air flow for breaths at the same temperature on
cooling and during euthermia. In hamsters, however, the flow rate
of the first breaths upon rewarming was significant less than the
flow rate of breaths taken during euthermia.
During rewarming, TB returned to euthermic levels in a linear
fashion in all age groups. Rates of rewarming were comparable
to rates of cooling, ranging from 0.37 ± 0.03 to 0.93 ± 0.11 ◦ C/min
(Tables 1 and 2). For those animals that successfully recovered from
respiratory arrest, breathing frequency increased with increasing
temperature, as did heart rate. With a few exceptions, both fR and
fH showed hysteresis when data from cooling were compared to
those for re-warming; during re-warming, at any given TB , fR and fH
were lower than at the same temperature during cooling (Figs. 1–4).
As during cooling, during warming to temperatures above roughly
20 ◦ C, breathing frequency was maintained while heart rate continued to rise.
4. Discussion
4.1. Developmental changes in neonatal cold tolerance
Early studies (Adolph, 1948a,b, 1951; Adolph et al., 1961;
Fairfield, 1948) comparing the cold tolerance of neonatal and
adult mammals established that neonatal mammals are more tolerant of low TB than adults and that neonatal cold tolerance is
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255
Fig. 6. The relationship between breathing frequency and heart rate during transitional cooling (filled circles) and subsequent re-warming (open circles) in neonatal and
juvenile (A) hamsters aged P2–3, P5–6, P8–9, P15–20, P26–28, P32–34, and (B) adult hamsters. Data are presented as absolute means ± SEM. Arrows indicate values during
cooling at 20 ◦ C.
gradually lost over development. Cold tolerance in these studies
was defined as the lower lethal temperature, the temperature at
which the heart failed. Adolph (1948a) and Fairfield (1948) reported
that the hearts of young rats and hamsters (various ages between
0 and 30 days of age) usually stopped beating at a TB just below
that at which breathing movements ceased. Subsequent studies
confirmed this difference in the temperatures at which breathing
and heart beat fail, and demonstrated that during apnea in deep
hypothermia (TB = 2–3 ◦ C) the hearts of neonatal mice (P0–10) continued to beat in the absence of breathing and that these animals
survived for prolonged periods provided that the glottis remained
open (Hill, 2000; Hill and Eshuis, 1988). This was attributed to gas
exchange due to pumping via cardiac oscillations and diffusion
and convection through an open glottis with the oxygen transport to tissues being maintained by the continuing heartbeat. In
the current study, the heart also continued to beat at a very low frequency in all neonatal and juvenile rats and hamsters for the 10 min
we waited following respiratory arrest at the minimum TB where
breathing failed. Of note is that respiratory arrest need not lead to
death (or even hypoxia) and that the temperature at which breathing ceases is not the same as the lower lethal temperature and
does not necessarily reflect the full extent of cold tolerance of the
animal.
In the present study, we found that the temperature at which
respiration ceased in neonatal rats during cooling was similar in
all age groups (ranging from 8.3 ± 1.7 to 10.7 ± 0.24 ◦ C) and did not
increase with age. None-the-less, it was lower than the temperature
at which breathing fails in adult rats (Adolph, 1951; Osborne and
Milsom, 1993). This implies that the mechanisms maintaining respiration at low temperatures do not change throughout this period
of development (P0–20) even though the mechanisms maintaining
the heartbeat (and establishing the lower lethal temperature) do.
Throughout the first two weeks of life the temperature at which the
heart fails increases (Adolph, 1951) approaching the temperature
at which breathing ceases.
In this context it is interesting to note that in the youngest
animals, both breathing frequency and heart rate fall in concert
but as the animals age, breathing frequency is maintained while
heart rate continues to fall during cooling at temperatures above
roughly 20 ◦ C. This suggests that during development, a degree of
temperature compensation occurs in the neural circuits controlling
breathing but not those controlling heart rate.
In the neonatal hamsters, respiratory cold tolerance actually increased during early development. The youngest neonates
investigated (P2–3) stopped breathing at 8.5 ± 0.7 ◦ C, while older
neonates and juveniles kept breathing at even lower temperatures.
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Fig. 7. TB of last breath before respiratory arrest during cooling and first breath after arrest during re-warming (autoresuscitation) for (A) rats aged P0–6, P14, P16, P18–20,
and (B) hamsters aged P2–3, P5–6, P8–9, P15–20, P26–28, P32–34, and adult. Data are presented as absolute means ± SEM. *Denotes a significant difference from the TB of last
breath before respiratory arrest (P ≤ 0.05, paired t-test). Note that this statistical test was not performed on rats aged P14, P16 and P18–20, and hamsters aged P32–34 and
P71–73 due to inability to resuscitate resulting in mortality. Letters are used to denote differences in the effect of hamster age on mean TB of arrest and of autoresuscitation:
bars with similar letters are not significantly different from each other, bars with different letters are (P ≤ 0.05, one-way repeated measures ANOVA). Sample sizes are
indicated in each bar. Data for rats aged P2–6 were obtained from Tattersall and Milsom (2003).
As hamsters grew, this respiratory cold tolerance was lost again
as animals aged. P32–34 hamsters arrested breathing at warmer
temperatures, similar to the TB of respiratory arrest in P2–3 pups.
4.2. Developmental changes in the ability to autoresuscitate
Adolph (1951) also observed that the age at which the lethal TB
of neonates reached that of the adult was a good predictor of the
age at which the ability to restart breathing spontaneously during warming was lost (∼19 days of age in rats and ∼30 days of
age in hamsters). Our data for neonatal rats is consistent with this
observation. In the current study the ability to autoresuscitate from
hypothermic respiratory arrest was lost in rats between P14 and
P20. In hamsters, however, the TB at which breathing restarted progressively fell throughout life and breathing only failed to restart
in animals that also underwent cardiac arrest. It was, however, at
P32–34 that we first observed individuals that also underwent cardiac arrest and were unable to recover from respiratory arrest. As
long as the heart kept beating, however, hamsters retained the
ability to autoresuscitate regardless of age. The heart rate during
respiratory arrest varied greatly between individuals and there did
not appear to be a difference between the fH of those individuals
that were destined to autoresuscitate upon rewarming after 10 min
and those in which fH and fR were destined to fail.
This initial increase in respiratory cold tolerance in hamsters
within the neonatal period has also been observed in vitro (Zimmer
and Milsom, 2004). The temperature at which breathing arrested
(i.e. neural respiratory discharge ceased) in 0–3 day old hamster brainstem spinal cord preparations was significantly higher
than that of 4–6 day old preparations. However, hamster pups
of equivalent age stopped breathing at much lower temperatures (∼4–8.5 ◦ C) than the temperature where the fictive discharge
stopped in vitro (∼14–15 ◦ C). Since the reduced preparation is
less cold tolerant than whole animals, something else must be
contributing to the persistent respiration at lower temperatures
in vivo. Presumably this arises from excitatory influences from
higher brain centers not present in the reduced, isolated brainstem
preparations.
Fig. 8. Tidal volume (VT ) and inspiratory time (TINSP ) of the first breath upon re-warming, a breath taken at the equivalent TB during cooling, and a breath taken at 35 ◦ C for
(A) rats aged P14–20 (n = 4), and (B) hamsters aged P26–28 (n = 4). VT is normalized as % of VT at 35 ◦ C. Air flow rate for each inspiration is indicated by the slope of the line.
Results are presented as mean values ± SEM. *Indicates a significant difference in slope from breath at 35 ◦ C (P ≤ 0.05, one-way repeated measures ANOVA on ranks).
A.E. Corcoran et al. / Respiratory Physiology & Neurobiology 181 (2012) 249–258
In summary, in rats the mechanisms maintaining respiration at
low temperatures did not change throughout development, rather
it was the processes that initiate breathing after respiratory arrest
that were altered (i.e. developmental changes in respiratory cold
tolerance and the ability to spontaneously restart breathing on
rewarming are not linked). In hamsters respiratory cold tolerance
increased throughout development and as long as the heart beat
was sustained, even adults recovered spontaneously from respiratory arrest.
Interestingly, there are similar parallels in developmental
changes in hypoxic responses in rats and hamsters. Both species
respond to hypoxia in similar fashion as neonates but their
responses to both acute and chronic hypoxia differ as adults
(Frappell and Mortola, 1994; Mortola, 1991).
4.3. Recovery is not via a gasping mechanism
It was reported earlier that gasping does not appear to play
a role in autoresuscitation from hypothermia-induced respiratory arrest in neonatal rats (Tattersall and Milsom, 2003). This
was not unexpected since there is considerable evidence to indicate that neonatal mammals do not suffer from hypoxia during
hypothermia-induced respiratory arrest (Hill, 2000). This was also
true in the present study. The first breath after re-warming was significantly longer, but not significantly larger than breaths taken at
the same temperature during cooling. As a result, the air flow rate
(as assessed by the slope of VT /TI ) of the first breath upon rewarming
was significantly lower than that of a normal breath indicating that,
rather than gasps, these breaths were more like sighs. Thus, our data
support the observation that gasping in neonatal and juvenile rats
and hamsters does not appear to play a role in autoresuscitation
upon re-warming. It also indirectly suggests that O2 uptake and
transport were maintained during the periods of respiratory arrest
and that the animals did not become anoxic.
4.4. Adaptive significance of cold tolerance and autoresuscitation
for young mammals
While the timeframe over which the ability to autoresuscitate
is lost does not correspond to the sensitive period in respiratory
development shown by others around P10–12 (Wong-Riley and Liu,
2008), there does appear to be a correlation between the ability
to recover from hypothermic respiratory arrest and thermoregulatory capacity. The loss of the ability to autoresuscitate in rats
and hamsters occurs as juveniles develop adult type homeothermy.
Prior to 15–17 days of age neither rats nor hamsters are capable of
maintaining TB in room temperature air (Fairfield, 1948; Sokoloff
et al., 2000; Spiers and Adair, 1986). By ∼20 days of age in rats
and ∼28 days in hamsters, pups are able to thermoregulate about
as well as adults, despite being much smaller (∼30% of their adult
weight). Thus the ability to autoresuscitate is lost in these animals
at approximately the same point in development that they gain
the ability to thermoregulate and avoid low TB . These changes in
thermoregulatory capacity are also consistent with the differences
seen in cold tolerance. Rat neonates were less cold tolerant and
showed no change over the time frame studied here while hamster neonates showed a better cold tolerance that increased early
in development. In this regard, neonatal rats exhibit moderate thermogenesis and are efficient huddlers (Blumberg, 1997; Sokoloff
and Blumberg, 2002). Neonatal hamsters, on the other hand, lack
endothermy during their first 12 post-natal days, after which they
begin to exhibit brown adipose tissue and shivering thermogenesis (Blumberg, 1997). Huddling behaviour often helps to maintain
body temperature, however this is rarely effective in hamsters
257
during exposure to cold until after the first week of life (Sokoloff
and Blumberg, 2002).
5. Conclusions
In summary, hypothermia in developing rats and hamsters led
to a progressive slowing of heart rate while breathing frequency
was roughly maintained down to a TB of ∼20 ◦ C. Beyond this point
fR also fell progressively. Cold tolerance, defined here as the TB at
which breathing arrested/restarted did not change in rat neonates
over the period studied while the ability to autoresuscitate from
hypothermic respiratory arrest was lost in rats between P14 and
P20. These data indicate that in rats, the mechanisms maintaining respiration down to low temperature were not changing, but
the processes that initiate breathing during rewarming was. This
further suggests that the loss of cold tolerance and the ability to
autoresuscitate in rats are not linked. In contrast, the cold tolerance
of hamsters increased until P26–28 while the ability to autoresuscitate was retained into adulthood, though continued heartbeat was
imperative for successful autoresuscitation. Additionally, hamsters
were consistently more cold tolerant than rats. Gasping did not
appear to play a role in autoresuscitation suggesting that severe
hypoxia does not develop during 10 min of hypothermic respiratory
arrest in juvenile rats and hamsters.
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