“…work with nonhuman primates forms a critical middle ground for future investigations of…neuro behavioral function after…anesthesia exposure…”

Image: John Ursino, ImagePower Productions.

Image: John Ursino, ImagePower Productions.

A SEMINAL report by Jevtovic-Todorovic et al.1  described neurotoxicity and long-term cognitive impairments after exposure to general anesthesia in infant (postnatal day 7) rats. This phenomenon has been observed repeatedly in multiple species and with a variety of different anesthetic drugs. It has also sparked parallel retrospective and prospective studies in humans that suggest that pediatric anesthesia might be associated with increased risk of adverse neurocognitive outcomes when exposure is repeated or prolonged.2–9  The recent investigation by Coleman et al.10  joins a growing number of studies investigating the impact of exposure to general anesthesia in infancy on neurobehavioral development in nonhuman primates. This highly translationally relevant model makes unique contributions to understanding the phenomena and potential mechanisms of long-term cognitive and behavioral changes after general anesthesia early in life.

There are a number of important advantages to studying this question in nonhuman primate models, not the least of which is the ability to study the effects of general anesthesia in the absence of surgery. Physiologic monitoring and support is much more feasible in an infant monkey than in an infant rodent. Thus, long-term effects of anesthesia exposure cannot easily be attributed to hypoxia, hypercapnia, and so forth. The stage of brain development at birth of a rhesus monkey is similar to that of a 6-month-old human infant,11,12  whereas based on many neurobiologic markers, postnatal day 7 rats that are at peak vulnerability to anesthetic neurotoxicity13  may more closely match the stage of brain development of a third-trimester human fetus. Nonhuman primates also have the capacity to perform more complex tasks and have a sophisticated social structure, like that of humans, allowing more translationally relevant tests of cognition and socioemotional behavior. Because puberty occurs between 3 and 5 yr of age in rhesus monkeys, it is possible to examine changes in behavior over the course of development that would be challenging to study in rodent models, in which sexual maturity occurs around the age of 6 weeks.

The primary outcomes in the study by Coleman et al.10  are measures of early motor reflexes and emotional behavior in a number of different situations. They employ test batteries for monkeys that were developed to be analogous to behavioral testing procedures in humans, including the Laboratory Temperament Assessment Battery14  and human intruder tests. These tests have been used extensively in studies of socioemotional development in macaque monkeys15  and therefore are well-validated and highly translational tools. Indeed, the use of these paradigms is a significant strength of studies in nonhuman primates where tests can be given in similar formats to humans and monkeys. As another example, the ongoing Mayo Anesthesia Safety in Kids study16  includes an operant behavioral testing battery that has been validated for developmental research in both humans and monkeys.17 

Coleman et al.10  found abnormal motor reflexes at 1 month of age (about 3 weeks after anesthesia exposure), as well as heightened anxiety in the home-cage social environment at 12 months of age in monkeys that were exposed three times to isoflurane (for 5 h each time) between postnatal days 6 and 12. Changes in these measures were not statistically significant in monkeys exposed to isoflurane only once for 5 h on postnatal day 6. Because monkeys did not undergo surgery, changes in anxiety in monkeys cannot be attributed to the experience of postoperative pain. These findings suggest that negative behavioral changes in children after surgery with general anesthesia18  might result, in part, from long-term effects of multiple exposures to the anesthetic agent during the surgical procedure.

Because of the time and expense involved in a prospective study with nonhuman primates, investigators have based their anesthesia protocols on durations of exposure that are known to result in increased neuro- and glioapoptosis,12,17,19,20  a candidate mechanism for the long-term effects of pediatric anesthesia on behavior. Because these durations tend to be longer than those commonly encountered in the pediatric operating room, many of these studies face the criticism that the anesthesia exposures are not clinically relevant. Single anesthesia exposures in infants that have been investigated recently8,9  are of short duration relative to those in the studies to date with monkeys. It is worth noting that prolonged anesthesia exposure is certainly not unknown in pediatric surgery; Coleman et al.10  cite 30% of infant anesthesia exposures being longer than 3 h at their institution, and prolonged sedation in the neonatal intensive care unit may last for weeks.17  However, repeated exposures to anesthesia appear rare in these populations; for example, 7.4% of cases (44/593) in the study by Wilder et al.4  received three or more anesthetic exposures, so three long exposures to anesthesia, especially within a week, would be extremely unusual in a clinical context. Although this is a limitation of extant preclinical studies, these exposure protocols establish boundary conditions upon which future work can build, to carry out more finely grained analyses of duration and frequency of anesthesia exposure.

Another issue in preclinical studies, especially with nonhuman primates, is that of repeated testing with a relatively small subject pool. This is a practical constraint of research with rare and expensive animals. Although this can incur statistical concerns about a large number of tests on a limited population, it is also a strength of these studies that data can be collected on the same study population longitudinally and across a number of different behavioral domains. As multiple research groups carry out studies using similar behavioral protocols, the generality of individual findings becomes clearer, as in all other areas of biomedical research. Similarly, this study like others tests both male and female monkeys but is underpowered to detect subtle sex differences. Nevertheless, this is a more desirable state of affairs than limiting preclinical studies to a single sex to evade this criticism.21 

Two studies in monkeys10,22  have independently reported increased anxiety-related behaviors in monkeys that were repeatedly exposed to general anesthesia as infants despite using different anesthetic agents and different schedules and durations of exposure. The finding that repeated exposure is associated with adverse neurocognitive outcomes is consistent with some human epidemiologic studies.4,23  Recent prospective studies in humans have emphasized the safety of single, relatively brief exposures to general anesthesia in the context of pediatric surgery8,9 ; Coleman et al.10  also find limited effects of single exposures to general anesthesia in monkeys. These observations reinforce one another and suggest that concerns about repeated or prolonged exposure of children to general anesthesia remain significant. Long-term neuro-behavioral changes after repeated exposure to anesthesia in infancy may not simply be a consequence of cumulative exposure to the anesthetic agent; one study in rodents found a greater decline in synaptic density in adult rats exposed to sevoflurane three times for 2 h each as infants compared to those exposed once for 6 h,24  so the mechanisms of long-term effects of repeated anesthesia exposure also merit further investigation.

The translation of these findings to the clinical setting remains subject to interpretation. It appears relatively easier to detect neurocognitive impairments in animal models after pediatric anesthesia than it does in clinical studies with humans. This may reflect uncertainty about the animal models that are used, with regard to details of anesthesia exposure, dosing, and so forth. It may also reflect the greater degree of experimental control in animal studies where all subjects receive identical anesthetic regimens, and behavioral assessments may be more extensive and sensitive to subtle effects.

Findings that single exposures to general anesthesia for pediatric surgery are not associated with adverse neurocognitive outcomes8,9  are a great relief to anxious parents (and anesthesiologists). However, it does not seem safe to conclude that because single exposures to general anesthesia for pediatric surgery are apparently without substantial long-term adverse neurocognitive effects, any pattern of pediatric anesthesia exposure will be similarly benign. This work with nonhuman primates forms a critical middle ground for future investigations of impaired neurobehavioral function after repeated and/or prolonged anesthesia exposure and for testing potential interventions to mitigate them.

Research Support

Supported in part by grant R01-HD068388 from the National Institutes of Health, Bethesda, Maryland.

Competing Interests

The authors are not supported by, nor maintain any financial interest in, any commercial activity that may be associated with the topic of this article.

References

1.
Jevtovic-Todorovic
V
,
Hartman
RE
,
Izumi
Y
,
Benshoff
ND
,
Dikranian
K
,
Zorumski
CF
,
Olney
JW
,
Wozniak
DF
:
Early exposure to common anesthetic agents causes widespread neurodegeneration in the developing rat brain and persistent learning deficits.
J Neurosci
2003
;
23
:
876
82
2.
O’Leary
JD
,
Janus
M
,
Duku
E
,
Wijeysundera
DN
,
To
T
,
Li
P
,
Maynes
JT
,
Crawford
MW
:
A population-based study evaluating the association between surgery in early life and child development at primary school entry.
Anesthesiology
2016
;
125
:
272
9
3.
Graham
MR
,
Brownell
M
,
Chateau
DG
,
Dragan
RD
,
Burchill
C
,
Fransoo
RR
:
Neurodevelopmental assessment in kindergarten in children exposed to general anesthesia before the age of 4 years: A retrospective matched cohort study.
Anesthesiology
2016
;
125
:
667
77
4.
Wilder
RT
,
Flick
RP
,
Sprung
J
,
Katusic
SK
,
Barbaresi
WJ
,
Mickelson
C
,
Gleich
SJ
,
Schroeder
DR
,
Weaver
AL
,
Warner
DO
:
Early exposure to anesthesia and learning disabilities in a population-based birth cohort.
Anesthesiology
2009
;
110
:
796
804
5.
Sprung
J
,
Flick
RP
,
Katusic
SK
,
Colligan
RC
,
Barbaresi
WJ
,
Bojanić
K
,
Welch
TL
,
Olson
MD
,
Hanson
AC
,
Schroeder
DR
,
Wilder
RT
,
Warner
DO
:
Attention-deficit/hyperactivity disorder after early exposure to procedures requiring general anesthesia.
Mayo Clin Proc
2012
;
87
:
120
9
6.
Ing
CH
,
DiMaggio
CJ
,
Malacova
E
,
Whitehouse
AJ
,
Hegarty
MK
,
Feng
T
,
Brady
JE
,
von Ungern-Sternberg
BS
,
Davidson
AJ
,
Wall
MM
,
Wood
AJ
,
Li
G
,
Sun
LS
:
Comparative analysis of outcome measures used in examining neurodevelopmental effects of early childhood anesthesia exposure.
Anesthesiology
2014
;
120
:
1319
32
7.
Ing
C
,
DiMaggio
C
,
Whitehouse
A
,
Hegarty
MK
,
Brady
J
,
von Ungern-Sternberg
BS
,
Davidson
A
,
Wood
AJ
,
Li
G
,
Sun
LS
:
Long-term differences in language and cognitive function after childhood exposure to anesthesia.
Pediatrics
2012
;
130
:
e476
85
8.
Sun
LS
,
Li
G
,
Miller
TL
,
Salorio
C
,
Byrne
MW
,
Bellinger
DC
,
Ing
C
,
Park
R
,
Radcliffe
J
,
Hays
SR
,
DiMaggio
CJ
,
Cooper
TJ
,
Rauh
V
,
Maxwell
LG
,
Youn
A
,
McGowan
FX
:
Association between a single general anesthesia exposure before age 36 months and neurocognitive outcomes in later childhood.
JAMA
2016
;
315
:
2312
20
9.
Davidson
AJ
,
Disma
N
,
de Graaff
JC
,
Withington
DE
,
Dorris
L
,
Bell
G
,
Stargatt
R
,
Bellinger
DC
,
Schuster
T
,
Arnup
SJ
,
Hardy
P
,
Hunt
RW
,
Takagi
MJ
,
Giribaldi
G
,
Hartmann
PL
,
Salvo
I
,
Morton
NS
,
von Ungern Sternberg
BS
,
Locatelli
BG
,
Wilton
N
,
Lynn
A
,
Thomas
JJ
,
Polaner
D
,
Bagshaw
O
,
Szmuk
P
,
Absalom
AR
,
Frawley
G
,
Berde
C
,
Ormond
GD
,
Marmor
J
,
McCann
ME
;
GAS Consortium
:
Neurodevelopmental outcome at 2 years of age after general anaesthesia and awake-regional anaesthesia in infancy (GAS): An international multicentre, randomised controlled trial.
Lancet
2016
;
387
:
239
50
10.
Coleman
K
,
Robertson
ND
,
Dissen
GA
,
Neuringer
MD
,
Martin
LD
,
Cuzon Carlson
VC
,
Kroenke
C
,
Fair
D
,
Brambrink
A
:
Isoflurane anesthesia has long-term consequences on motor and behavioral development in infant rhesus macaques.
Anesthesiology
2017
;
126
:
74
84
11.
Dobbing
J
,
Sands
J
:
Comparative aspects of the brain growth spurt.
Early Hum Dev
1979
;
3
:
79
83
12.
Brambrink
AM
,
Evers
AS
,
Avidan
MS
,
Farber
NB
,
Smith
DJ
,
Zhang
X
,
Dissen
GA
,
Creeley
CE
,
Olney
JW
:
Isoflurane-induced neuroapoptosis in the neonatal rhesus macaque brain.
Anesthesiology
2010
;
112
:
834
41
13.
Yon
JH
,
Daniel-Johnson
J
,
Carter
LB
,
Jevtovic-Todorovic
V
:
Anesthesia induces neuronal cell death in the developing rat brain via the intrinsic and extrinsic apoptotic pathways.
Neuroscience
2005
;
135
:
815
27
14.
Goldsmith
HH
,
Rothbart
MK
:
Contemporary instruments for assessing early temperament by questionnaire and in the laboratory
in
Explorations in Temperament
. Edited by
Angleitner
A
,
Strelau
J
.
New York, Springer
,
1991
, pp
249
72
15.
Kalin
NH
,
Shelton
SE
:
Defensive behaviors in infant rhesus monkeys: Environmental cues and neurochemical regulation.
Science
1989
;
243
:
1718
21
16.
Gleich
SJ
,
Flick
R
,
Hu
D
,
Zaccariello
MJ
,
Colligan
RC
,
Katusic
SK
,
Schroeder
DR
,
Hanson
A
,
Buenvenida
S
,
Wilder
RT
,
Sprung
J
,
Voigt
RG
,
Paule
MG
,
Chelonis
JJ
,
Warner
DO
:
Neurodevelopment of children exposed to anesthesia: Design of the Mayo Anesthesia Safety in Kids (MASK) study.
Contemp Clin Trials
2015
;
41
:
45
54
17.
Paule
MG
,
Li
M
,
Allen
RR
,
Liu
F
,
Zou
X
,
Hotchkiss
C
,
Hanig
JP
,
Patterson
TA
,
Slikker
W
Jr
,
Wang
C
:
Ketamine anesthesia during the first week of life can cause long-lasting cognitive deficits in rhesus monkeys.
Neurotoxicol Teratol
2011
;
33
:
220
30
18.
Stargatt
R
,
Davidson
AJ
,
Huang
GH
,
Czarnecki
C
,
Gibson
MA
,
Stewart
SA
,
Jamsen
K
:
A cohort study of the incidence and risk factors for negative behavior changes in children after general anesthesia.
Paediatr Anaesth
2006
;
16
:
846
59
19.
Brambrink
AM
,
Back
SA
,
Riddle
A
,
Gong
X
,
Moravec
MD
,
Dissen
GA
,
Creeley
CE
,
Dikranian
KT
,
Olney
JW
:
Isoflurane-induced apoptosis of oligodendrocytes in the neonatal primate brain.
Ann Neurol
2012
;
72
:
525
35
20.
Slikker
W
Jr
,
Zou
X
,
Hotchkiss
CE
,
Divine
RL
,
Sadovova
N
,
Twaddle
NC
,
Doerge
DR
,
Scallet
AC
,
Patterson
TA
,
Hanig
JP
,
Paule
MG
,
Wang
C
:
Ketamine-induced neuronal cell death in the perinatal rhesus monkey.
Toxicol Sci
2007
;
98
:
145
58
21.
Clayton
JA
,
Collins
FS
:
Policy: NIH to balance sex in cell and animal studies.
Nature
2014
;
509
:
282
3
22.
Raper
J
,
Alvarado
MC
,
Murphy
KL
,
Baxter
MG
:
Multiple anesthetic exposure in infant monkeys alters emotional reactivity to an acute stressor.
Anesthesiology
2015
;
123
:
1084
92
23.
Flick
RP
,
Katusic
SK
,
Colligan
RC
,
Wilder
RT
,
Voigt
RG
,
Olson
MD
,
Sprung
J
,
Weaver
AL
,
Schroeder
DR
,
Warner
DO
:
Cognitive and behavioral outcomes after early exposure to anesthesia and surgery.
Pediatrics
2011
;
128
:
e1053
61
24.
Amrock
LG
,
Starner
ML
,
Murphy
KL
,
Baxter
MG
:
Long-term effects of single or multiple neonatal sevoflurane exposures on rat hippocampal ultrastructure.
Anesthesiology
2015
;
122
:
87
95