1. Introduction
The presence of an affiliative conspecific (or cues associated with a
conspecific) reduces stress responses to a wide variety of stimuli
(Davitz & Mason, 1955; Lyons et al. , 1988; da Costa et
al. , 2004; Bowen et al. , 2013; Kanitz et al. , 2014). This
phenomenon is known as “social buffering” (Kiyokawa, 2017; 2018;
Kiyokawa & Hennessy, 2018). Numerous studies in a variety of species
have demonstrated that, in addition to social buffering by the mother or
mate (Hennessy et al. , 2009; Smith & Wang, 2014), social
buffering can be elicited by other conspecifics (Lyons et al. ,
1993; Winslow et al. , 2003; Kiyokawa et al. , 2004;
Hennessy et al. , 2008). Given that these three classes of
conspecifics induce social buffering via distinct neural mechanisms,
specification of the type of conspecific is important when discussing
the neural mechanisms underlying social buffering (Kiyokawa & Hennessy,
2018).
We demonstrated in rats that social buffering induced by a conspecific
other than the mother or mate can ameliorate stress responses to an
aversively conditioned stimulus (CS). When a fear-conditioned rat was
exposed to the CS alone, conditioned fear responses, including increased
freezing and hypothalamic-pituitary-adrenal axis activation, were
observed. However, the presence of a non-conditioned unfamiliar same-sex
rat (associate) completely blocked these responses (Kiyokawa et
al. , 2004; Kiyokawa et al. , 2007; Kiyokawa & Takeuchi, 2017).
Subsequent studies revealed a number of characteristics (Kiyokawaet al. , 2004; Ishii et al. , 2016; Mikami et al. ,
2016; Kiyokawa et al. , 2018; Kiyokawa et al. , 2019; Mikamiet al. , 2020) and a possible neural pathway underlying social
buffering of conditioned fear responses (Kiyokawa et al. , 2009;
Kiyokawa et al. , 2012; Takahashi et al. , 2013; Kiyokawaet al. , 2014; Fuzzo et al. , 2015; Minami et al. ,
2019). However, little information is available regarding the
neurochemical background of social buffering.
We propose opioids, dopamine, oxytocin, and vasopressin as candidates
responsible for social buffering because they play an important role in
affiliative behavior toward conspecifics other than the mother or mate.
For example, play behavior between same-sex adolescent rats is reduced
by treatment with an opioid receptor antagonist (Beatty & Costello,
1982). Another study using social play as an unconditioned stimulus
demonstrated that opioid receptor antagonists block the establishment of
conditioned place preference (Trezza et al. , 2011). Dopamine is
an additional candidate because treatment with a dopamine receptor
antagonist was shown to reduce intrinsic play behavior between same-sex
adolescent rats (Beatty et al. , 1984; Niesink & Van Ree, 1989)
and optogenetically induced social investigation between female mice
(Gunaydin et al. , 2014). In addition to opioids and dopamine,
oxytocin and vasopressin should be included among the list of candidates
based on their established roles in the formation of attachment to one’s
children and mates. For example, after giving birth, ewes exhibit
rejection behavior toward alien lambs. However, oxytocin infusion
reverses such rejection behavior (Kendrick et al. , 1992).
Similarly, in monogamous prairie voles, males and females show a
preference for their mates over unfamiliar opposite-sex conspecifics.
However, this partner preference is blocked by treatment with
vasopressin antagonists in males (Winslow et al. , 1993) and
treatment with oxytocin antagonists in females (Insel & Hulihan, 1995).
Consistent with these findings, an oxytocin antagonist was found to
block social buffering induced by bonded males in female prairie voles
(Adam, 2014). In addition, some studies have suggested possible roles
for oxytocin and vasopressin in affiliative behavior toward same-sex
conspecifics. For example, treatment with an oxytocin antagonist was
shown to reduce intermale social investigation both in rats and mice
(Lukas et al. , 2011). A similar effect was observed in
vasopressin gene–knock-down male mice (Rigney et al. , 2022).
Based on these findings, we hypothesized that antagonists of one or more
of these neurochemicals would block social buffering of conditioned fear
responses in rats.
To elucidate the neurochemical background of social buffering, it is
important to identify the sites at which responsible neurochemicals act.
The presence of opioids (Stein, 2016) or dopamine (Einhorn et
al. , 1991) decreases the intrinsic excitability of neurons, whereas the
presence of oxytocin (Rogers-Carter et al. , 2018) or vasopressin
(Liu et al. , 2003) increases the intrinsic excitability of
neurons. Therefore, during antagonist-mediated blockade of social
buffering, the antagonist would modulate neural activity at the site of
action within the neural pathway related to social buffering and/or
affiliative behavior. One methodological approach for identifying the
site of modulation would be to observe the changes in Fos expression
throughout the brain. We expect that Fos expression would be altered at
the sites at which responsible neurochemicals act.
In this study, a series of experiments were performed to explore the
neurochemical background of social buffering in rats. In Experiment 1,
fear-conditioned subject rats were first injected intraperitoneally with
either naloxone (a non-selective opioid receptor antagonist),
haloperidol (a dopamine D2 receptor antagonist), atosiban (a oxytocin
receptor antagonist), SR49059 (a vasopressin V1a receptor antagonist),
or saline. The subjects were then exposed to the CS with an associate.
The effect of each antagonist was evaluated by observing thebehavioral
responses. Experiment 2 examined the effect of naloxone on locomotor
activity during an open-field test. In Experiment 3, we analyzed Fos
expression in 16 brain regions of the second cohort of subjects during
naloxone-mediated blockade of social buffering.