Department of Veterinary
Sciences, University of Pisa, Italy
Queen Mother Hospital for
Small Animals, Royal Veterinary
College,
Hertfordshire,
UK
Abstract
Maternal care has been shown to affect the
development of the brain, behaviour, social
skills and emotional systems of the young of
many mammalian species including dogs. The aim
of the present study was to determine the
effects of maternal care on the behavioural
responses of family dog puppies towards
environmental and social stimuli. In order to do
this, maternal care (licking puppy's ano-genital
area, licking the puppy, nursing and
mother-puppy contact) during the first three
weeks after birth was assessed in 12 litters of
domestic dog puppies reared in home environments
(total = 72 puppies). The behavioural responses
of puppies were assessed in an arena and an
isolation test, which were performed when the
puppies were two-month old. Data were analysed
using principal components analysis and
projection to latent structures regression. A
systematic relationship was found between
maternal care and behaviour in both tests. In
the arena test, maternal care was found to be
positively associated with approach to the
stranger, attention oriented to the stranger,
time spent near the enclosure, yawning, whining
and yelping (R2Y = 0.613, p = 8.2 _ 10_9).
Amount of maternal care was negatively
associated with the number of squares crossed
and the time spent individually playing with the
rope. In the isolation test, the amount of
maternal care was positively associated with
standing posture, paw lifting, and howling, and
it was negatively associated with yawning, lying
down and nose licking (R2Y = 0.507, p =
0.000626). These results suggest that the amount
of maternal care received during early life
influences the pattern of behavioural responses
and coping strategies of puppies at two-months
of age. On the basis of these findings it could
be speculated that early maternal care
contributes to adaption to the environment in
which family puppies are developing, with
particular regard to social relationships with
people.
Résumé
Il a été
démontré que les soins maternels
affectent le développement du cerveau, du
comportement, des aptitudes sociales et des
systèmes émotionnels des
progénitures de nombreuses espèces
de mammifères, y compris les chiens.
Le but de la présente étude
était de déterminer les effets des
soins maternels sur les réponses
comportementales des chiots aux stimuli
environnementaux et sociaux. Pour ce faire, les
soins maternels (lécher le chiot, les
soins et le contact mère-chiot) ont
été évalués chez 12
portées de chiots domestiques
élevés dans des environnements
domestiques (total = 72 chiots).
Les réponses comportementales des
chiots ont été
évaluées dans une pièce
ouverte et au cours d'un test d'isolement, qui
ont été effectuées lorsque
les chiots étaient âgés de
deux mois.
Une relation a été
trouvée entre les soins maternels et le
comportement dans les deux tests. Dans le test
de la pièce ouverte, les soins maternels
se sont positivement associés à
l'approche de l'étranger, à
l'attention portée à
l'étranger, au temps passé
près de la niche, aux bâillements,
aux gémissements et aux jappements.
Dans le test d'isolement, la quantité
de soins maternels était positivement
associée à la posture debout, au
soulèvement des pattes et aux hurlements,
et elle était négativement
associée au bâillement, au coucher
et au léchage du nez.
Ces résultats suggèrent que la
quantité de soins maternels reçus
au début de la vie influence le
modèle des réponses
comportementales et des stratégies
d'adaptation des chiots à l'âge de
deux mois. Sur la base de ces résultats,
on pourrait supposer que les soins maternels
précoces contribuent à
l'adaptation à l'environnement dans
lequel se développent les chiots
domestiques, en particulier en ce qui concerne
les relations sociales avec les personnes.
Introduction
Maternal care plays a crucial role in the
life of the offspring; in most mammals it
ensures neonatal survival and represents the
main source of stimuli in the early postnatal
environment. Its influence on neuro-behavioural
development has been well documented in a
variety of species [1,2,3] including
rodents [4,5,6,7], non-human primates
[8,9], and humans [10,11].
In mice and rats there is a connection
between maternal grooming and ano-genital
licking received by pups in the early postnatal
period, and their later behavioural patterns
[12], including those related to stress
responses, fear and anxiety [1,13].
Highly responsive maternal behaviour is known to
promote a stress neurobiology that is less
reactive as well as more resilient to challenge.
The mechanisms are highly specific and involve
relatively permanent modifications of DNA that
control the expression of glucocorticoid
receptors [14].
In monkeys, such as the Geoffroy's marmoset
[9] and Rhesus macaque [15], low
quality or abusive maternal care during the
early postnatal period produces more pronounced
hypothalamic-pituitary adrenal (HPA) responses
to environmental stressors throughout
development and into adulthood, compared with
animals reared with higher quality early
maternal care.
In human infants, sensitive and responsive
maternal caregiving has an equivalent role in
buffering the HPA system [16,17], and a
higher quality of maternal behaviour has been
found to predict a better cortisol recovery
after a mildly stressful event in three-month
old infants [18].
With respect to domestic dogs, the study of
maternal behaviour has previously received
comparatively little attention [3].
Rheingold's landmark text on maternal behaviour
in dogs [19] has only recently been
followed up by other peer reviewed works in this
area [3,20,21,22,23,24,25]. With regard
to the effects of maternal care on the
behavioural development of puppies, and on the
subsequent behaviour of adult dogs, only few
works based on observational methods have been
published [26,27,28]. Results have also
been inconsistent. Foyer and colleagues
[26] found that, in female military
German shepherd dogs, there was individual
variation in the expression of maternal
behaviour. They also found that mother-offspring
interactions were associated with individual
differences in physical and social engagement,
and aggression, in the offspring. Guardini et
al. [27] studied the effects of morning
maternal care on 8-week-old puppies living in
standardised rearing conditions with a low level
of socialisation towards people. This study
showed that, as previously found in rodents (see
[6,7]), the quantity of maternal care
received by puppies reared in standardised
conditions during the first three weeks of life
mediates a set of responses which allows the
individual to cope with stressful situations and
to better adapt to the environment. Similarly,
Tiira & Lohi [29] found, through the
use of a questionnaire administered to
dog-owners, that fearfulness in family dogs was
associated with poorer maternal care during
puppyhood. However, in contradiction to the
findings of Foyer et al. [26] and
Guardini et al. [27], Bray et al.
[28] found that increased maternal
behaviour was positively associated with
undesirable anxiety-related behaviours and
performance in young adult dogs.
To our knowledge, the effects of maternal
care on the response of family reared puppies
towards unfamiliar environmental and social
stimuli has not previously been investigated.
This was the focus of the present study.
The offspring-directed maternal behaviour of
12 mothers, from a range of breeds, was recorded
during the first three weeks after the birth of
the puppies. The behaviour of these puppies was
then assessed at two months of age in two
potentially stressful situations (arena and
isolation tests).
In a previous study we applied the same
methodology to a population of laboratory dogs
[27]. In that study it was found that a
higher level of maternal care was associated
with more exploratory and fewer anxiety related
behaviours during the isolation test. No
systematic relationship was found between
maternal care and arena test behaviour. However,
human socialisation and external stimulation
provided to laboratory puppies were
significantly lower than for family reared
puppies.
We therefore hypothesised that family
puppies would behave differently from laboratory
puppies in those situations in which social
(unfamiliar person) and inanimate stimu 4.
Discussion
Early life experiences are known to shape
the behavioural development of animals. Previous
studies in different species have demonstrated
that maternal care plays a key role in the
offspring's ontogeny (e.g., [1,3,31]),
and such results have also been found in
domestic dogs [26,27,28]. However, to
our knowledge, this is the first study
investigating the effect of maternal care on the
behaviour of two-month old puppies that have
been reared in a family environment.
In the present study, the use of
multivariate statistical methods (PLS and OPLS)
identified statistically significant systematic
relationships between maternal care and
behaviour in both the arena and isolation tests.
This suggests that the amount of maternal care
(in terms of nursing, body licking, ano-genital
licking, and physical contact between mother and
puppy) received during the 21 days after birth
has an effect on the pattern of behaviour of
family-reared puppies when they are faced with
unfamiliar environmental and social stimuli at
two months of age. As opposed to individual
correlations between variables, multivariate
relationships of this kind indicate the presence
of patterns of altered behaviours that point to
an underlying latent process.
The puppies' behaviour in the arena test was
systematically associated with the amount of
maternal care received during the first 21 days
of life (as summarised by the maternal care
score). The strongest positive loadings
(variables positively associated with maternal
care score) within that model were certain
stress behaviours (increase of yawning and
whining/yelping), the time spent near the
enclosure and the puppies' interest in the
unfamiliar human figure (increased orientation
and approach). In contrast, the strongest
negative loadings (variables negatively
associated with maternal care score) were the
puppies' activity, interest in his/her
surroundings and the inanimate stimuli (reduced
number of crossed squares and time spent playing
with the rope). Increased maternal care was
associated with increased social interest shown
by family reared puppies towards the stranger in
the arena. The pattern of positive loadings
found in the arena test in the present study are
comparable with those behaviours observed during
the activation of the attachment system when a
young mammal is separated from its attachment
figure in an unfamiliar environment, as observed
in the Ainsworth Strange Situation Test (ASST,
[10]), which is a paradigm created for
the assessment of the attachment bond between
children and their caregivers. The stress signs
and the vocalisations emitted by the family
reared puppies in the arena test are comparable
with the protest behaviours children show when
separated from their mothers [10], which
have also been documented in infant monkeys
[32,33], lambs [34], and
domestic dog puppies [35,36,37].
Remaining near the enclosure boundary could
be interpreted as an attempt to regain proximity
to the attachment figure. This behaviour is
similar to "stay by" and "be oriented to the
door" in children [10] and adult dogs
[38] when tested in the ASST, and to the
attempts to break the barriers separating infant
rhesus monkeys from their mothers [39].
It seems that the more maternal care a family
reared puppy receives during early life, the
more distress he/she shows during separation,
and the more orientated he/she is to
reunification with the attachment figure or
obtaining social support from another social
partner (in this case the stranger in the
arena). This is likely to be due to the higher
quality of the attachment bond between the
mother and the offspring. The puppies stay close
to the unfamiliar person, and attempt to seek
attention in order to obtain comfort and support
in the absence of the attachment figure. The
same tendency to seek contact and support from
an unfamiliar individual, in the absence of a
familiar individual or attachment figure, has
been previously observed in children
[11], young chimpanzees [40],
and adult dogs [38,41,42]. In the
present study, the level of maternal care seems
to influence the puppy's motivation and/or
strategy when faced with a stressful situation,
in this case seeking support and comfort from an
unfamiliar human being. The puppies' attempt to
buffer stress through social contact with a
human is supported by the work of Pettijhon et
al. [43], in which it was observed that
the human figure was very effective in
alleviating separation-induced distress
vocalisation in young puppies, especially if the
person behaved actively rather than passively.
In the current study, the stranger was
instructed to behave passively, and therefore
any relief from separation distress that the
puppies might have obtained from contact with
that person would only have been partial. Stress
behaviours observed in the arena test in the
present study might therefore also have been
influenced by this failed distress
buffering.
The fact that maternal care has an impact
upon the puppies' interest in a social stimulus
is an important finding, and is in agreement
with studies in other mammalian species (e.g.,
rats: [44,45]; prairie voles:
[46]; rhesus macaques: [47,48],
including humans [49] and adult military
German shepherds dogs [26]. With regard
to an underlying biological basis for this
effect on social behaviour, it has been observed
that the quantity and the quality of maternal
care influences oxytocin receptor density in the
brain in rodents [2,44], humans
[31,50], in rhesus monkeys [51],
as well as various neurochemicals which
including vasopressin, prolactin,
catecholamines, endogenous opioids,
adrenocorticotropic hormone, and
gamma-aminobutyric acid [52]. Although
there is a lack of knowledge about the impact of
maternal care upon oxytocin levels in Canis
Familiaris puppies, it is likely that a similar
mechanism is present in this species.
The tendency to seek security from strangers
is potentially very important for domestic dog
puppies, who have to forge a close relationship
with humans when they are taken away from their
mothers and introduced into a new home. A
positive attitude towards human strangers at two
months of age, which is the usual time of
adoption, may in fact make the separation from
the mother and the establishment of a bond with
the new owners easier. Further research is
needed in order to more deeply understand the
link between the amount of maternal care,
oxytocin levels and puppies' attitude towards
people.
In our previous study [27], the same
protocol was used to investigate the influence
of morning maternal care on two-month old
puppies reared in standardised conditions with
limited exposure to human beings. The findings
were quite different: In that study, no
significant systematic relationship was found
between maternal care score and the puppies'
behaviour in the arena test. This difference may
be due to general effects of maternal care on
the puppy's adaptability to the environment and
his/her inclination to seek social support,
combined with the higher level of interspecific
socialisation of the family reared puppies that
enabled them to regard an unfamiliar person as a
potential source of security. The presence of
this source of security may also have enabled
the puppies to orientate their coping strategy
more consistently, leading to a more systematic
organisation of behaviour that was not
detectable in laboratory puppies.
Contrary to the original hypothesis, higher
scores for maternal care were associated with
reduced interest in the environment or the
objects that were present in the arena test.
These findings also contradict some of the
previous literature on the effects of maternal
care in other species. In rodents, increased
ano-genital licking and grooming by the mother
is correlated with an increase in play behaviour
by the offspring [31,53,54], as well as
in exploratory behaviour [4,5,13]. This
could, again, be explained as being a
consequence of the developmental environment;
puppies reared in a family are likely to be more
focussed on people than on physical stimuli
during the arena test. This explanation is
supported by data from a previous work
[27], which found that higher morning
maternal care scores were associated with
increased exploratory behaviour in puppies
reared in an environment with low interspecific
socialisation.
The puppies' behaviour in the isolation test
was also systematically associated with the
amount of maternal care received during the
first 21 days of life, although this model was
weaker than the arena test behaviour model. The
strongest positive loadings in this model were
for standing posture, paw lifting and howling.
Howling is a distress vocalisation, which can be
interpreted as et-epimeletic or care soliciting
behaviour [35,55,56], emitted by the
puppies in order to call the mother and to
regain contact with her. Standing posture could
be interpreted as the puppy waiting for signs of
social reunification (i.e., with the mother). As
in the arena test, it seems that the more
maternal care a puppy received during his/her
early life, the greater his/her need to regain
proximity with his/her mother during separation
(and hence the greater the distress observed
during isolation). The important difference
between the arena and the isolation test was the
absence, in the latter, of any form of potential
social support. When family puppies were
completely alone they vocalised, but if a
stranger was present, puppies tried (although
unsuccessfully) to seek comfort and support from
that person (otherwise they showed separation
distress). It can be hypothesised that the level
of maternal care influences the type of stress
responses shown by family puppies in this
stressful situation (absence of the mother and
littermates and being in an unfamiliar
environment). In the isolation test model, the
strongest negative loadings were with lying,
yawn, and nose licking. The latter two are
subtle stress signs [57].
The initial hypothesis that we would find
similar results in the isolation test to those
of our previous study involving laboratory
reared puppies [27] was not confirmed.
In the laboratory puppies, the amount of
maternal care was most strongly positively
associated with engagement with the environment
inside the metal fences, less destructive
behaviours and reduced non-exploratory
locomotion. Those results are more similar to
those from the literature in rodents (see
[1,4,5,6,13,58,59,60]). The difference
may relate to the diversity of breeds in the
present study, and the different sensory
environments to which the puppies were exposed.
A family home is a much more complex and
stimulating environment than a laboratory
kennel, and the family reared puppies had far
greater contact with people. It is possible that
family reared puppies found the arena
environment less engaging because they were
already used to more complex and stimulating
environments. The opposite could be said for the
laboratory puppies.
Another potentially influential factor in
this study is litter size. Litter size can have
an effect on the amount of maternal behaviour
available to individual puppies; when the
mother's time and energy are divided between
members of a larger litter, we might expect
individual mother-pup interactions to be
reduced. Unfortunately, due to the small number
of litters and relatively uniform litter size in
the present study, it was not possible to study
the effect of litter size. Given the confounding
effects of environmental variation within and
between homes, if the effect of litter size were
to be investigated, it would probably be easier
to identify an effect in laboratory reared pups
with a standardised environment.
The results of the present study are likely
to be more applicable to dogs that live in an
enriched environment that includes social
stimuli, as is the experience of most companion
and working dogs. Interestingly, Bray et al.
[28] also found an association between
the amount of maternal care received and anxiety
behaviours in young guide dogs between 14 and 17
months of age; dogs that received higher levels
of maternal care showed behaviours related to
stress and anxiety when isolated in an empty
room (e.g., high activity and short latency to
vocalisations when presented with a novel
object). Bray et al. [28] took into
account the nursing style of the mothers and
found that mothers whose nursing style (vertical
nursing) required greater effort by puppies were
more likely to produce offspring that were
successful in cognitive and temperament tasks,
whereas mothers that used a ventral nursing
style that required less effort from the puppies
were more likely to produce offspring that
failed. The authors proposed that there might be
benefits from a moderate amount of stress during
early life: vertical nursing style may provide
opportunities for puppies to cope with small
challenges which are beneficial and adaptive in
the long term, enhancing arousal regulation and
resilience.
Future research may clarify how maternal
factors, the social environment and also genetic
effects contribute to the expression of pet
dogs' behaviour in different phases of
development, from puppyhood to adulthood.
5. Conclusions
The quantity of maternal care received by
family puppies in early life influences their
behaviour in an unknown environment, in the
presence and in the absence of a stranger.
Specifically, maternal care was associated not
only with increased interest in a human
stranger, but also with increased display of
separation-related stress. Although these
findings disagree with some previous studies of
the development of animals, including our own
study involving puppies reared in conditions of
low socialisation with humans, they help to
provide further insight into the subtle
interaction between maternal behaviour and
environmental exposure during development.
