DOI: 10.5061/dryad.5mkkwh78c
title: Cooperative burrowing in mice

description: Animals often adjust their behavior according to social context, but the
 capacity for such behavioral flexibility can vary among species. Here, we
 test for interspecific variation in behavioral flexibility by comparing
 burrowing behavior across three species of deer mice
 (genus Peromyscus) with divergent social systems, ranging from
 promiscuous (P. leucopus and P. maniculatus) to
 monogamous (P. polionotus). First, we compared the burrows built by
 individual mice to those built by pairs of mice in all three species.
 While burrow length did not differ in P.
 leucopus or P. maniculatus, we found
 that P. polionotus pairs cooperatively constructed
 burrows that were nearly twice as long as those built by individuals
 and that opposite-sex pairs dug longer burrows than same-sex
 pairs. Second, to directly observe cooperative digging behavior
 in P. polionotus, we designed a burrowing assay in which we could
 video-record active digging in narrow, transparent enclosures. Using this
 novel assay, we found, unexpectedly, that neither males nor females spent
 more time digging with an opposite-sex partner. Rather, we demonstrate
 that opposite-sex pairs are more socially cohesive and thus more efficient
 digging partners than same-sex pairs. Together, our study demonstrates how
 social context can modulate innate behavior and offers insight into how
 differences in behavioral flexibility may evolve among closely related
 species.
description: <em>Large
 enclosures.<strong> </strong></em>We measured
 burrow architecture as described previously (Weber & Hoekstra
 2009; Weber<em> et al.</em> 2013; Metz<em> et
 al.</em> 2017). Briefly, at the start of the dark cycle, we
 introduced individuals or pairs of mice into large PVC boxes (1.2 x 1.5 x
 1.1m) filled with approximately 700kg hydrated, hard-packed premium play
 sand (Quickrete, Atlanta, GA). In the enclosures, mice were provided food
 and water <em>ad libitum</em> along with a cotton nestlet. We
 removed mice from the enclosures after one (pregnancy trials) or two
 (standard trials) overnight periods. Then, we made casts of the resultant
 burrows using polyurethane filling foam (Hilti, Schaan, Liechtenstein).
 Next, a researcher blind to trial identity hand-measured the burrow casts.
 For each trial, the number of burrows and total length of each burrow was
 recorded. We also categorized burrow shape as either “simple” or “complex”
 as described previously (Weber & Hoekstra 2009): <em>P.
 leucopus</em>and <em>P. maniculatus</em> burrows are
 considered “simple” with a short entrance tunnel and terminal nest
 chamber, whereas <em>P. polionotus</em> burrows are considered
 “complex” with a long entrance tunnel, nest chamber, and escape tunnel.
 After each trial, we removed all food, feces, nesting material, and
 disturbed substrate from the enclosure, and then rinsed the enclosure
 walls with water and turned over the sand to minimize residual
 odors.  <em>Narrow, transparent
 enclosures.</em> We also assayed burrowing behavior of
 mice directly using an acrylic chamber (5cm x 90 cm x 60 cm) with a
 transparent Plexiglass face (see Fig. 3A). Using a pre-cut mold, we
 sculpted hydrated sand into two symmetrical 45° hills and excavated an 8cm
 tunnel from one randomly selected hill to encourage burrowing in a
 consistent location. We outfitted the apparatus with an infrared
 illuminator frame that enabled video recording in the dark. We then
 introduced individuals or pairs of mice into an enclosure at the start of
 the dark cycle and removed animals the next morning, recording 8 hours of
 continuous video during the dark cycle. Mice were provided food and
 water <em>ad libitum</em>. Photographs of the resultant
 burrows were taken at the end of the trial. Details of photo and video
 analyses are provided below. Following each trial, we removed all food,
 feces, and disturbed substrate and rinsed the enclosure to minimize
 residual odors.