Tom 68 2019 Numer 4 (325) Strony 561–574

Ewa Joanna Godzińska, Julita Korczyńska, Anna Szczuka

Laboratory of Ethology Nencki Institute of Experimental Biology PAS 3 Pasteur Str., 02-093 Warsaw E-mail: [email protected]

DYADIC NESTMATE REUNION TEST IN THE RESEARCH ON SOCIAL BEHAVIOR

INTRODUCTION AS CONVENIENT SUBJECTS FOR COMPARATIVE RESEARCH ON BIOLOGICAL ETHOLOGY: AIMS, SCOPE AND METHODS ROOTS OF SOCIAL BEHAVIOR One of the most important behavioral Comparative research devoted to biolog- sciences, ethology, can be defined as the ical roots of social behavior owes a lot to study of mechanisms and evolution of be- numerous studies using as subjects social havior, focused both on accurate observa- , in particular the honeybee (Apis tion and description of behavior and on mellifera) and many of ants (Wilson its causation. Ethology is concerned with 1975; Godzińska 2005, 2006, 2016, 2019; four main categories of causal factors of Gowdy and Krall 2016; Moffett 2018). behavior: (1) immediate causal factors, The use of ants as subjects in behavioral acting „here and now”, (2) factors that research provides many advantages in com- had acted in the course of the ontogeny parison with the use of the honeybee. Ants of the studied individuals, (3) factors that are much more long-lived than honeybees: had acted in the course of the phylogeny ant workers may live up to several years, of the studied species, and, lastly, (4) fac- and ant queens up to 30 years (Höll- tors related to function (adaptive signifi- dobler and Wilson 1990; Keller and Ge- cance) of the studied behavioral trait. The noud 1997; Keller 1998; Godzińska et al. first two categories and the second two 1999; Jemielity et al. 2005). Long lifespan ones are designated jointly, respectively, of ants facilitates long-term research and as proximate and ultimate (evolutionary) experiments devoted to ontogeny and ac- causal factors of behavior. Causation of quired modifications of behavior (Godzińska behavior is studied mainly by means of 1997). Behavioral repertories of ants are experiments carried out both in laboratory very rich, and social behavior of these in- conditions and in the field with the use of sects shows great complexity, plasticity and various behavioral bioassays. In contrast flexibility H ( ölldobler and Wilson 1990, to some other behavioral sciences, general 1998, 2009; Godzińska 1996a, 2007, 2016; theoretical concepts of ethology arose as Korczyńska 1996; Szczuka 1996; Wnuk a result of comparative studies of many 2013; Mazurkiewicz et al. 2015). Richness phylogenetically distant groups, in- and complexity of ant behavior makes pos- cluding not only vertebrates, but also in- sible comparative analysis of social behavior of ants and humans (Wilson 1975; Jaisson vertebrates (Tinbergen 1963; Chmurzyński 1993; Godzińska 2005, 2016; Wnuk 2013; 1973; Thorpe 1979; Godzińska 1996b, 1997, 2019). Gowdy and Krall 2016; Moffett 2018). In particular, ants were used as model organ-

Keywords: ants, behavior, dyadic nestmate reunion test, Formicidae, social behavior Acknowledgements: This work was supported by the Statutable Grant 407 of the Ministry of Science and Higher Educa- tion for the Nencki Institute of Experimental Biology of the Polish Academy of Sciences in Warsaw, Poland. 562 Ewa Joanna Godzińska i współaut. isms in comparative behavioral and neu- test may be preceded by various treatments robiological research on biological roots of applied to one or both tested individuals, aggression and violence (Kostowski 1994; such as social isolation, food deprivation, Hölldobler and Wilson 1998; Godzińska and various acute or chronic pharmacologi- 2005, 2007; Mazurkiewicz et al. 2015), and cal treatments (Wnuk and Godzińska 2006). in the experiments investigating pro-social behavior, and, in particular, rescue behavior DYADIC NESTMATE REUNION TESTS (coming to the rescue of endangered indi- PRECEDED BY SOCIAL ISOLATION AND viduals) (Czechowski et al. 2002; Nowbaha- ISOLATION-INDUCED TROPHALLAXIS ri and Hollis 2010; Hollis and Nowbahari Dyadic nestmate reunion tests were 2013a). Ant behavior also shows astound- used in numerous studies investigating the ing diversity, which is not surprising, as behavior of ants reunited with a nestmate several thousands of valid extant ant spe- after a period of social isolation. Ants were cies have already been described on a world usually isolated singly in test tubes. At the scale, and many other species remain still start of each test two tubes were opened undiscovered. More than one hundred ant and their open ends were put together, one species were found to inhabit Poland (Höll- against the other, to allow a confrontation dobler and Wilson 1990; Czechowski et al. of a dyad of ants (Boulay et al. 1999a, b, 2012; Mazurkiewicz et al. 2015). Last but 2000b, 2004; Cybulska et al. 2000; Kat- not least, ant behavior can be conveniently zav-Gozansky et al. 2004; Korczyńska et studied in both laboratory and field condi- al. 2005; Wagner-Ziemka et al. 2006, 2008; tions (Hölldobler and Wilson 1990, 1998, Mazurkiewicz et al. 2016). 2009; Godzińska 1996a, 1997, 2006, 2007; The authors of the first studies investi- Wnuk and Godzińska 2006; Mazurkiewicz gating the behavior of ants during dyadic et al. 2015). In particular, ants show inter- nestmate reunion tests carried out after a esting behavioral and physiological respon- period of social isolation used as subjects ses to modifications of social contextW ( il- carpenter ants from the Campono- son 1971; Szczuka 1996; Godzińska 1997, tus, namely Camponotus fellah (Boulay et 2006, 2019; Wnuk and Godzińska 2006; al. 1999a, b, 2000b, 2004; Katzav-Gozan- Wnuk 2013; Mazurkiewicz et al. 2015; see sky et al. 2004), Camponotus herculeanus also Szczuka et al. 2019, this issue of KO- (Korczyńska et al. 2005), and two African SMOS). species, Camponotus acvapimensis (Cybul- The aim of the present paper is to show ska et al. 2000) and Camponotus maculatus1 how a relatively simple behavioral bioassay (Wagner-Ziemka et al. 2008; Mazurkiewicz et may provide a wealth of important find- al. 2016). These studies discovered, among ings and contribute to the extension of our others, the phenomenon of the so called knowledge about the behavior of the tested isolation-induced trophallaxis. Carpenter species, its causation and its variability. We ant workers reunited with a nestmate af- choose the so called dyadic nestmate reun- ter a period of social isolation ranging from ion test as an example of such a simple 24 hours (Camponotus fellah: Boulay et al. and yet very useful bioassay. 1999b; Camponotus acvapimensis: Cybulska et al. 2000) to 20 days (Camponotus fellah: DYADIC NESTMATE REUNION TEST Boulay and Lenoir 2001; Camponotus her- culeanus: Korczyńska et al. 2005) showed increased propensity to engage in trophal- GENERAL DESCRIPTION laxis, a specific category of social contacts Dyadic nestmate reunion test consists of widespread in societies. Trophallaxis a confrontation of two nestmates (members most frequently involves mutual contacts of the same colony or social group) placed of mouthparts of two individuals accompa- together in the same container. Behavior nied by the exchange of liquid food and/or of both individuals is recorded and then various active compounds, in particular cu- analyzed, usually in a detailed way. Ana- ticular hydrocarbons playing a crucial role lyzed behavior patterns may include vari- in the mediation of nestmate recognition ous aggressive and non-aggressive contacts (Hölldobler and Wilson 1990, 1998, 2009; between the tested ants, various forms of Cybulska and Godzińska 1999; Godzińska self-focused behavior such as self-grooming, 2006; Mazurkiewicz et al. 2015; Leonhardt various types of resting behavior and loco- 1 motion, and various behavioral responses The ants used in these two studies (Wagner-Ziemka et al. 2008, Mazurkiewicz et al. 2016) were first identifed as to elements of physical environment present Camponotus melanocnemis (Bolton, pers. comm.), and that during the test. Video recording of the whole species name was used in the paper of Wagner-Ziemka et test makes possible very accurate identifica- al. (2008). However, that species name is now invalid, and, tion of all specific behavior patterns. The therefore, the correct name (Camponotus maculatus) has been used in the paper of Mazurkiewicz et al. (2016). Dyadic nestmate reunion test in the research on ant social behavior 563 et al. 2016; see also Szczuka et al. 2019, of neuroethological methods involving ab- this issue of KOSMOS). Cuticular hydro- dominal injections of various neuroactive carbons are synthesized by each ant and compounds (Boulay et al. 1999a, 2000b; then exchanged during trophallaxis, allog- Korczyńska et al. 2005). These experiments rooming (= licking another individual) and revealed that isolation-induced trophallaxis other social contacts. They are then stored can be partly suppressed by the administra- in the post-pharyngeal glands located in the tion of relatively high doses of the biogen- worker heads, and still later are deposited ic amine octopamine (Boulay et al. 1999a; on the body surface of the ants during self- Korczyńska et al. 2005; see also Szczuka et grooming. As a consequence of these ex- al. 2019, this issue of KOSMOS). Octopa- changes ant nestmates bear similar cuticu- mine, a biogenic amine widespread in inver- lar hydrocarbon profiles on their body sur- tebrates and often considered to represent face. That phenomenon plays a crucial role the functional equivalent of noradrenaline, in nestmate recognition (Godzińska 2006; acts as a neurotransmitter, neuromodulator Wnuk and Godzińska 2006; Mazurkiewicz and neurohormone, and participates in the et al. 2015; Leonhardt et al. 2016; see also mediation of many aspects of insect physiol- Szczuka et al. 2019, this issue of KOSMOS). ogy and behavior (Roeder 2003; Roeder et Isolation-induced trophallaxis was ob- al. 2005) including the phenomena of posi- served not only during dyadic nestmate re- tive reinforcement and reward involved in union tests (Boulay et al. 1999a, b, 2000b, the associative olfactory learning of the hon- 2004; Cybulska et al. 2000; Katzav-Gozan- eybees (Hammer and Menzel 1998; Farooqui sky et al. 2004; Korczyńska et al. 2005; et al. 2003; Giurfa and Sandoz 2012). It Wagner-Ziemka et al. 2008; Mazurkiewicz is thus possible that octopamine injections et al. 2016), but also in several other stud- reduced isolation-induced trophallaxis by ies during which isolated ants were rein- mimicking rewarding effects of trophallactic troduced into their maternal nest (Boulay exchanges between nestmates. However, the et al. 2000a), a colony fragment created by phenomenon of octopamine-mediated social splitting their maternal colony into smaller reward accompanying ant trophallactic be- units (Boulay et al. 2004), or an experimen- havior remains hypothetical (Korczyńska et tal arena containing a large group of their al. 2005; see also Szczuka et al. 2019, this nestmates (Boulay et al. 2003; Katzav-Go- issue of KOSMOS). zansky et al. 2004). An alternative hypothesis concerning Behavior of carpenter ants during dy- causal factors responsible for inhibitory ef- adic nestmate reunion tests carried out af- fects of octopamine administration on isola- ter a period of social isolation was shown tion-induced trophallaxis takes into account to depend in a very important way on the the results of two experiments that demon- duration of that period. In particular, ants strated that administration of octopamine that had been subjected to relatively short may reduce not only behavioral consequenc- social isolation period (1-10 days) were usu- es of complete social deprivation (social iso- ally not attacked by their nestmates during lation), but also behavioral modifications dyadic nestmate reunion tests (Cybulska et induced by various forms of partial social al. 2000; Boulay et al. 2004; Korczyńska deprivation (Szczuka and Godzińska 2008; et al. 2005) or when reintroduced to their Vander Meer et al. 2008; see also Szczuka maternal nest (Boulay et al. 2000a). How- et al. 2019, this issue of KOSMOS). In the ever, longer isolation periods (14-40 days) study of Vander Meer et al. (2008) workers induced aggressive responses of nestmates of the fire ant Solenopsis invicta responded of the tested ants as a consequence of more to queen removal by significant decrease important divergence of cuticular hydrocar- of both brain octopamine levels and nest- bon profiles between isolated and non-iso- mate recognition acuity that manifested it- lated ants (Boulay et al. 2000a, 2003, 2004; self as decreased aggressiveness towards Katzav-Gozansky et al. 2004). However, this non-nestmate conspecifics. However, chronic rule was not held unexceptionally: in the oral octopamine treatment counterbalanced study of Korczyńska et al. (2005) workers of both these effects. Chronic oral adminis- C. herculeanus did not engage in aggressive tration of octopamine was also shown to behavior during dyadic nestmate reunion reduce modifications of responses to dead tests irrespectively of the duration of social insect prey induced by dramatic reduction isolation period (5 or 20 days). of worker group size in workers of the red wood ant Formica polyctena (Szczuka and NEUROCHEMICAL ASPECTS OF ISOLATION- Godzińska 2008). As demonstrated by ear- INDUCED TROPHALLAXIS lier research of Szczuka and Godzińska The phenomenon of the isolation-induced (1997, 2000, 2004a, b), ants of that species trophallaxis was also investigated by means kept in relatively small worker groups cease 564 Ewa Joanna Godzińska i współaut. to respond to dead insect prey by perform- havior and metabolism exerting antagonistic ing complete sequences of predatory/scav- effects with respect to octopamine (Roeder enging behavior terminated by prey retrieval 2003; Roeder et al. 2005). In the discussed to the nest. This behavioral modification is, study of Korczyńska et al. (2005) behavio- however, reversible and may be reduced or ral effects of octopamine and tyramine were even completely disappear if workers kept never antagonistic, and were even strikingly in a small group (about 25 individuals) are similar in the case of several analyzed be- subjected to chronic oral administration of havioral categories. However, only octopa- octopamine (Szczuka and Godzińska 2008). mine administration influenced significantly All these data taken together (Boulay et al. both trophallaxis and other social contacts 1999b, 2000b; Korczyńska et al. 2005; Szc- observed during that experiment. Tyramine zuka and Godzińska 2008; Vander Meer et did not exert any significant effects on ant al. 2008) suggest strongly that octopamine social behavior. reduces in ants various behavioral effects of various forms of social deprivation: complete BEHAVIOR OF NON-TROPHALLACTING ANTS social deprivation, queen deprivation and re- DURING DYADIC NESTMATE REUNION TESTS duction of worker group size. CARRIED OUT AFTER SOCIAL ISOLATION However, yet another study devoted to Trophallaxis is encountered in the ma- complex interrelationships between social jority of ant species, but not in all ants isolation, trophallaxis, and octopamine levels (Hölldobler and Wilson 1990; Cybulska in ant brains (Wada-Katsumata et al. 2011) and Godzińska 1999; Mazurkiewicz et al. brought about different results. In that 2015). In particular, trophallaxis is absent study workers of Formica japonica that had in some myrmicine ants from arid , been subjected to 2 day social isolation had including harvester ants from the genera significantly higher brain octopamine levels Pogonomyrmex and Messor, and the ants of than both control (non-isolated) workers and the genus Aphaenogaster (Wilson and Eis- ants isolated for only 1 h. However, if ants ner 1957; Delage 1968; Delage and Jais- isolated during 2 days were allowed to in- son 1969; Lenoir et al. 2001a, b; Ichinose teract with two nestmates before the meas- et al. 2005). urements of their brain octopamine levels, Lenoir et al. (2001a) investigated the no significant differences were discovered behavior of non-trophallacting workers of between control ants, ants isolated during 1 Aphaenogaster senilis reintroduced to their h, and ants isolated during 2 days. In other mother nests after various periods of social words, brain octopamine levels of workers isolation (2, 4, 6, 8, 10, 15 and 20 days), of F. japonica increased as a consequence and the effects of duration of social isola- of social isolation, but decreased again as tion on profiles of cuticular hydrocarbons a consequence of social interactions with present on their body surface and in their nestmates involving trophallaxis and allo- post-pharyngeal glands. Both these cat- grooming. egories of cuticular hydrocarbon profiles Neurochemical treatments applied to car- showed progressive changes as a result of penter ants before the start of dyadic nest- social isolation. As a consequence, ants that mate reunion tests also included abdominal had been subjected to relatively short isola- injections of three other biogenic amines: tion periods (2-10 days) induced friendly so- serotonin (Boulay et al. 2000b), dopamine cial contacts (mainly allogrooming) in their (Korczyńska et al. 2005), and tyramine resident nestmates, but if they had been (Korczyńska et al. 2005). However, none of subjected to longer isolation periods (15-20 these compounds exerted any significant ef- days), they were attacked. This experiment fect either on trophallactic behavior, or on also revealed that in non-trophallacting other social contacts observed during dyadic workers of A. senilis allogrooming represents nestmate reunion tests. This was surprising, the major way of transfer of nestmate rec- as in the mediation of behavior of social in- ognition cues. That last conclusion was ful- sect serotonin often acts in an antagonistic ly confirmed by Lenoir et al. (2001b) who way with respect to octopamine (Erber et used radioactive tracers to study the trans- al. 1993; Pribbenow and Erber 1996). Both fer of radiolabeled cuticular hydrocarbons serotonin and dopamine are also known to between nestmates of A. senilis during both play a crucial role in the mediation of ant dyadic nestmate reunion tests and group aggressive behavior (Kostowski 1994; Szczu- tests consisting of a confrontation of a do- ka et al. 2013). Lastly, tyramine, a metabol- nor ant with 10 recipients. ic precursor of octopamine, also widespread Wagner-Ziemka et al. (2006) applied dy- in invertebrates and considered to repre- adic nestmate reunion tests carried out af- sent a functional equivalent of adrenaline, ter 3 days of social isolation to study in is also known to act as a modulator of be- detail the behavior of workers of A. senilis Dyadic nestmate reunion test in the research on ant social behavior 565 taken from two colonies separated by about 2011; Wnuk 2013; Korczyńska et al. 2014; 40 km. In this study, too, social contacts Bernadou et al. 2015; Symonowicz et al. observed during the tests involved mainly 2015; Dussutour et al. 2016; see also Sz- various forms of allogrooming and antennal czuka et al. 2019, this issue of KOSMOS). contacts. Allogrooming was, however, less Wagner-Ziemka et al. (2008) and Mazurk- frequent than expected and in the case of iewicz et al. (2016) compared the behavior one of the tested colonies was observed only of intranidal versus extranidal workers of during a half of the tests. Moreover, behav- the African species Campono- ior of the tested ants showed a surprisingly tus maculatus during dyadic nestmate re- large number of significant inter-colony dif- union tests carried out after 48 h of social ferences. Among others, ants from one of isolation. Categories of workers compared the tested colonies much more frequently in these experiments included minor nurses engaged in resting behavior and licking of and foragers (Wagner-Ziemka et al. 2008), the inner walls of the set of two test tubes and intranidal and extranidal major workers in which they were confined during the (Mazurkiewicz et al. 2016). All workers inves- test. That last behavior might have been tigated in these two studies were taken from triggered by chemical cues left there by in- the same colony. teracting nestmates. The behavior of minor nurses and minor foragers of C. maculatus showed numerous DYADIC NESTMATE REUNION TESTS IN THE differences. Nurses showed higher propensity RESEARCH ON BEHAVIORAL POLYMORPHISM to engage in some forms of social contacts AND DIVISION OF LABOR IN ANT COLONIES (mostly antennal contacts and trophallaxis Dyadic nestmate reunion tests were also and, to a lesser degree, also allogrooming) applied to study behavioral differences be- and in some forms of resting behavior. At tween nestmates from the same colony the same time, they showed lower propensity (Wagner-Ziemka et al. 2008; Mazurkiewicz to engage in behavior patterns displayed in et al. 2016: Camponotus maculatus, a high- response to various elements of their physi- ly polymorphic African carpenter ant spe- cal environment (Wagner-Ziemka et al. 2008). cies; Korczyńska et al. 2014: F. polyctena, a Similar differences between intranidal and monomorphic species belonging to the group extranidal workers were discovered also in of the red wood ants). Colonies of polymor- major workers of C. maculatus: intranidal phic ants contain minor workers (minors) majors showed higher propensity for resting with relatively small bodies and heads, ma- behavior and lower propensity for behavio- jor workers (majors) with large bodies and ral responses to elements of physical envi- disproportionately large heads, and some- ronment than extranidal ones. However, in times also intermediate forms, media workers contrast to minor nurses and foragers of C. (Hölldobler and Wilson 1990; Wilson 1971; maculatus, intranidal and extranidal majors Mazurkiewicz et al. 2015, 2016). However, in of that species did not differ with respect to colonies of both monomorphic and polymor- propensity for locomotion, self-grooming and phic ants also exists division of labor related social behavior. In particular, longer trophal- to worker age and/or degree of its behavioral lactic interactions had relatively high rate and physiological maturation, the so called of occurrence during the tests with minor age or temporal polyethism. Young workers nurses and both intranidal and extranidal engage first in inside-nest (intranidal) activi- majors, but much lower rate of occurrence ties and usually act as nurses taking care of during the tests with minor foragers. In oth- the queen(s) and developing brood. As they er words, the transition from intranidal to age, they switch to outside-nest (extranidal) extranidal tasks seems to be accompanied activities and become foragers (Wilson 1971; by decreased propensity for trophallaxis only Hölldobler and Wilson 1990, 2009; Wnuk in the case of minors of C. maculatus. Ma- 2013; Mazurkiewicz et al. 2015; see also jors of that species retain high propensity for Szczuka et al. 2019, this issue of KOSMOS). trophallaxis also after having switched to ex- However, foragers may return to intranidal tranidal tasks (Mazurkiewicz et al. 2016). activities and become the so called reverted Korczyńska et al. (2014) compared the nurses in response to modifications of social behavior of five categories of workers of the context such as exposure to brood in ab- red wood ant F. polyctena: callows (newly sence of younger workers acting as nurses. eclosed intranidal workers), nurses (relatively This process, the so called behavioral rever- young workers collected from the ant mound sion, was already extensively studied in vari- and then kept together with brood and col- ous ants (Ehrhardt 1931; Dobrzańska 1959; ony foragers), colony foragers (ants that had Lenoir 1979a, b; Sorensen et al. 1984; Mc- been collected from the ant mound and then Donald and Topoff 1985; Godzińska 2006; kept together with brood and nurses in lab- Wnuk and Godzińska 2006; Wnuk et al. oratory nests in which they acted as forag- 566 Ewa Joanna Godzińska i współaut. ers), trail foragers (collected from the trails DYADIC NESTMATE REUNION TESTS NOT and then kept in isolation from nurses and PRECEDED BY SOCIAL ISOLATION IN brood), and reverted nurses (workers that THE RESEARCH ON ANT TROPHALLACTIC had already switched to outside-nest activi- BEHAVIOR ties, but then returned back to the role of the nurse). The reverted nurses were experi- Dyadic nestmate reunion tests were also mentally created from trail foragers by ex- used to study ant trophallactic exchanges posing them to brood in absence of ordinary not induced by social isolation. Ants used nurses. Before the test all tested workers as subjects in that research belonged to the were subjected only to very short social iso- carpenter ant species lation (15 min) to avoid possible disruption (Bonavita-Cougourdan et al. 1979; Bonavi- of the process of behavioral reversion in the ta-Cougourdan and Gavioli 1981; Bonavita- reverted nurses as a consequence of their Cougourdan 1983; Bonavita-Cougourdan deprivation from contacts with brood. and Morel 1984a, b, 1985, 1986, 1988; Behavioral differences between five cate- Morel 1986). Two interacting workers ei- gories of ants investigated by Korczyńska et ther had the same behavioral specialization al. (2014) were related both to their age and (forager) (Bonavita-Cougourdan and Gavioli their past and present behavioral specializa- 1981; Bonavita-Cougourdan 1983; Bonavi- tion. Not surprisingly, the youngest ants, ta-Cougourdan and Morel 1986, 1988), or newly eclosed callows, were the least active. differed with respect to age (callow or ma- Nurses, relatively young workers that parti- ture worker) (Bonavita-Cougourdan and cipated still only in intranidal tasks, usually Morel 1984a, b, 1985, 1988; Morel 1986) behaved in a way intermediate with respect and/or behavioral specialization (nurse ver- to behavior of callows and the ants that had sus forager) (Bonavita-Cougourdan and Mo- already passed the transition to extranidal rel 1984a, b, 1985; Morel 1986). A large tasks. Reverted nurses behaved similarly as part of these experiments involved the anal- both classes of foragers and differently than ysis of antennal communication between nurses with respect to numerous behavior workers of C. vagus during trophallactic patterns ranging from immobility to agonis- contacts taking place during dyadic nest- tic behavior and non-aggressive social be- mate reunion tests (Bonavita-Cougourdan havior. However, behavioral variables quan- 1983; Bonavita-Cougourdan and Morel tifying many other traits of behavior of re- 1984a, b, 1985; Morel 1986). According verted nurses took intermediate values with to Lenoir (1982), antennal movements ac- respect to both nurses and foragers. Only a companying trophallactic exchanges between few traits of behavior were closely similar in workers of Myrmica rubra transmit much both ordinary and reverted nurses. This im- less information than some other communi- plies that behavioral reversion of workers of cation systems used by social insects, such F. polyctena does not consist of the return as honeybee dances. Ant antennal com- of foragers to fully nurse-like behavior. Last- munication cannot thus be involved in the ly, the behavior of colony and trail forag- transfer of precise, detailed information and ers showed both similarities and numerous acts rather as modulatory communication differences. This is not surprising, as two that can only mediate activity level of the groups of foragers differed with respect to partner ant. The results of the experiments origin (mound versus trails), present social with C. vagus supported fully that conclu- environment (presence vs absence of con- sion (Bonavita-Cougourdan 1983, Bonavita- tacts with nurses and brood), and probably Cougourdan and Morel 1984b). However, also worker age (trail foragers were presum- antennal movements accompanying trophal- ably older than colony foragers). laxis showed interesting differences related The results of that study also support to age and/or behavioral specialization of the hypothesis that honeybee and ant be- interacting workers (Bonavita-Cougourdan havioral reversion show qualitative differ- and Morel 1984a, b, 1985; Morel 1986). ences (Lenoir 1979a; Sorensen et al. 1984; In particular, the behavior of mature donor see also Szczuka et al. 2019, this issue of ants differed as a function of their behavio- KOSMOS). As shown by Huang and Robin- ral specialization (nurse versus forager), but son (1996), deprivation of honeybee foragers only during trophallactic exchanges with ma- from contacts with younger workers acting ture receivers. That effect was not observed as nurses may induce behavioral and mor- if a mature donor engaged in trophallaxis phological phenotype changes characteristic with an immature (callow) receiver. Moreo- for behavioral reversion. However, depriva- ver, during trophallactic exchanges with cal- tion from contacts with younger workers did lows antennal activity of mature donors was not result in the induction of nurse-like be- more variable and its organization was dif- havior in foragers of F. polyctena. ferent than during trophallaxis with mature Dyadic nestmate reunion test in the research on ant social behavior 567 receivers (Bonavita-Cougourdan and Morel during dyadic confrontations with four cat- 1984a, 1985; Morel 1986). egories of opponents: a nestmate, a non- Bonavita-Cougourdan et al. (1979) also nestmate conspecific ant, an allospecific ant invented an interesting method of analy- (a worker of Formica fusca), and a potential sis of liquid flow transmitted between two prey, a small nymph of the house cricket ants engaged in trophallaxis. That method (Acheta domesticus). As revealed by that ex- involves measurements of radioactivity emit- periment, during dyadic confrontations of ted by the radioactive isotope of gold (Au198) foragers of F. polyctena with nestmate or added to honey offered to the donor be- non-nestmate conspecifics the rate of occur- fore its dyadic encounter with the receiver. rence of specific behavior patterns showed The use of that method led, among others, very little differences irrespectively of the to a surprising discovery that the direction type of the opponent and neurochemical of food flow between two foragers of C. va- treatment received by the tested ant. In all gus may be repeatedly reversed during the these situations overt aggression was almost same act of trophallactic behavior. Work- absent, and interactions of two opponents ers that practised such food flow reversals involved mostly ritualized aggression (open- also showed higher propensity to engage in mandible threats) and friendly social behav- trophallaxis, and were more aggressive to- ior patterns including frequent trophallactic ward non-nestmate conspecifics (Bonavita- exchanges. Interestingly, octopamine, known Cougourdan and Gavioli 1981; Bonavita- to reduce isolation-induced trophallaxis Cougourdan and Morel 1988). (Boulay et al. 1999b, 2000a; Korczyńska et al. 2005), did not reduce trophallactic DYADIC NESTMATE REUNION behavior of workers of F. polyctena during TESTS IN THE RESEARCH ON ANT dyadic confrontations with conspecifics that AGGRESSIVE BEHAVIOR had not been preceded by social isolation. Other biogenic amines used in that study (dopamine, serotonin and tyramine) GENERAL REMARKS also did not exert any influence on trophal- Dyadic nestmate reunion tests were also lactic behavior observed during dyadic con- carried out as control tests in numerous frontations between nestmates. However, studies investigating ant aggressive behavior both serotonin and dopamine exerted in- by means of dyadic aggression tests. These hibitory effects on trophallactic contacts ob- studies yielded interesting results extending served during dyadic confrontations between our knowledge about aggressive behavior of non-nestmate conspecifics. ants from the genus Formica including For- Finally, all four amines used in that mica lugubris (Le Moli and Parmigiani 1982; study exerted inhibitory effects on allo- Le Moli et al. 1983b, 1984; Parmigiani and grooming observed during dyadic confron- Le Moli 1987), F. polyctena (Le Moli and tations with nestmates, and serotonin and Mori 1986; Parmigiani and Le Moli 1987; tyramine exerted inhibitory effects also on Beye et al. 1997; Szczuka et al. 2013), For- antennal contacts between nestmates (Szc- mica pratensis (Beye et al. 1998) and Formi- zuka et al. 2013; see also Szczuka et al. ca rufa (Le Moli et al. 1982, 1983b, 1984; 2019, this issue of KOSMOS). Parmigiani and Le Moli 1987; Kleineidam et al. 2017). Other ant species tested in these DYADIC NESTMATE RESCUE TESTS IN studies included two other formicine spe- THE RESEARCH ON ANT PRO-SOCIAL cies, Polyrhachis laboriosa (Mercier et al. BEHAVIOR 1997) and Cataglyphis iberica (Dahbi and Lenoir 1998), and myrmicine species Acro- ANT RESCUE BEHAVIOR myrmex octospinosus (Jutsum et al. 1979), Leptothorax nylanderi and Leptothorax gre- Pro-social behavior is defined as ac- dleri (Heinze et al. 1996), and A. senilis tions that are intended to benefit anoth- (Ichinose et al. 2005). er individual/other individuals (Bartal et al. 2011, 2014; Ueno et al. 2019). One of THE EFFECTS OF ADMINISTRATION OF the most interesting subcategories of risky BIOGENIC AMINES ON BEHAVIOR OF ANTS pro-social behavior has been called the DURING DYADIC NESTMATE REUNION TESTS rescue behavior (Czechowski et al. 2002). NOT PRECEDED BY SOCIAL ISOLATION Rescue behavior is defined as a social in- Szczuka et al. (2013) investigated behav- teraction during which one individual, the ioral consequences of acute administration victim, is endangered, and another indi- of four biogenic amines (dopamine, seroto- vidual, the rescuer, places itself at risk nin, octopamine and tyramine) on behavior of endangerment by engaging in rescue of workers of the red wood ant F. polyctena attempts. The behavior of the rescuer 568 Ewa Joanna Godzińska i współaut. should also be generally suited to the cir- DYADIC NESTMATE RESCUE TESTS IN THE cumstances, and should not be inherent- RESEARCH ON ANT RESCUE BEHAVIOR ly rewarding or beneficial to the rescuer Since several years dyadic nestmate res- (Nowbahari and Hollis 2010; Hollis and cue tests consisting of a confrontation of a Nowbahari 2013a). single freely moving ant with a single en- The research on ant rescue behavior dangered nestmate (captured by a predator was carried out both in the field and labo- or entrapped in an artificial snare) were also ratory conditions with the use of two main increasingly frequently used in the research bioassays: antlion larva capture bioassay, investigating various aspects of ant rescue in which rescue behavior of the tested ants behavior (Taylor et al. 2013; Miler 2016; is elicited by stimuli emitted by a victim Miler et al. 2017a, b; Miler and Kuszewska ant captured by a predatory antlion larva 2017). These studies documented many in- (Czechowski et al. 2002; Miler 2016; Mil- teresting phenomena. Among others, Taylor er et al. 2017a, b; Taylor et al. 2013), et al. (2013) found out that a single worker and artificial snare (entrapment) bioassay, of Tetramorium species E may successfully in which rescue behavior of potential res- rescue its nestmate captured by an antlion cuers is elicited by stimuli emitted by a larva. Taylor et al. (2013) also observed an victim ant entrapped in an artificial snare interesting sequence of events during which (Nowbahari et al. 2009, 2012, 2016; Hollis the victim ant was released by the antlion as and Nowbahari 2013b; Taylor et al. 2013; a consequence of a successful rescue action, Cichoń 2017; Duhoo et al. 2017; Kurasz- but then the rescuer was in turn grabbed kiewicz 2017; Miler et al. 2017a, b; Mil- by the predator, and the original victim, now er and Kuszewska 2017). Recently, Uy et freed, began to engage in rescue behavior. al. (2019) used a modified version of the This observation demonstrated that rescue entrapment bioassay in which victim ants, actions may be promptly reciprocated. workers of the weaver ant Oecophylla smar- A series of experiments of Miler and his agdina, were experimentally immobilized by coworkers explored various factors influenc- being wrapped in spider silk. Yet another ing nestmate rescue behavior of workers of type of ant rescue behavior was reported in F. cinerea during bioassays with antlion lar- the termite-hunting ant Megaponera ana- vae (Miler 2016) and artificial snares (Mil- lis: workers of that species were observed er and Kuszewska 2017), and during both to transport injured nestmates back to the these types of bioassays (Miler et al. 2017a, nest and to engage in their intense al- 2017b). These studies revealed, among oth- logrooming that facilitated wound healing ers, that workers of F. cinerea with life ex- (Frank et al. 2017, 2018). Workers of the pectancy experimentally shortened by expo- harvester ant Veromessor pergandeyi were sure to carbon dioxide less efficiently trig- also observed to free nestmates ensnared gered rescue behavior of their nestmates in spider webs, transport them to the nest, during the bioassays with antlion larvae and groom away their silk bindings (Kwapi- (Miler 2016), and less readily engaged in ch and Hölldobler 2019). rescue behavior in response to a nestmate Ant rescue behavior is most frequent- entrapped in an artificial snare (Miler et al. ly investigated by means of a test during 2017b). These findings were surprising, as which five freely moving ants act as po- in ants shortened life expectancy often leads tential rescuers of a nestmate entrapped to increased readiness to engage in risky in an artificial snare (bound to a piece of tasks (Moroń et al. 2008, 2012). Therefore, filter paper that helps to capture phero- it might have been expected that ants with mones emitted by the victim ant). Such shortened life expectancy will show higher tests were applied to study rescue be- propensity to engage in risky rescue behav- havior of workers of Cataglyphis cur- ior, However, many studies reported, that sor (Nowbahari et al. 2009, 2012, 2016; in ants shortened worker life expectancy Duhoo et al. 2017) and F. polyctena may also lead to social withdrawal (Höll- (Cichoń 2017; Kuraszkiewicz 2017). Arti- dobler and Wilson 1990; Korczyńska 2001; ficial snare bioassay was also used in the Ugelvig and Cremer 2007; Heinze and Wal- field, close to ant nests, to study rescue ter 2010; Bos et al. 2011; Diez et al. 2015; behavior of workers from several Mediter- Leclerc and Detrain 2017). The findings of ranean ant species (Cataglyphis floricola, Miler (2016) and Miler et al. (2017b) can Lasius grandis, Aphaenogaster senilis, thus be interpreted, too, in terms of the Messor barbarus and Messor marocanus) induction of social withdrawal. If so, these (Hollis and Nowbahari 2013b), and North findings suggest that experimental studies American ant species Tetramorium species of ant rescue behavior may prove to be of E and Prenolepis imparis (Taylor et al. importance for comparative research devoted 2013). to causal factors underlying social withdraw- Dyadic nestmate reunion test in the research on ant social behavior 569 al, and that they perhaps may even shed Absence of rescue behavior in behavioral light on biological roots of autism spectrum repertories of these three ant species was disorders. This last possibility is very in- most probably related to very low risk of triguing, as some autism-related genes were entrapment faced by these ants in their already found in the genome of the honey natural environment. , a social insect closely related to the ants (Shpigler et al. 2017). DYADIC REUNION TESTS AND DYADIC Dyadic nestmate rescue tests with the RESCUE TESTS IN BEHAVIORAL use of an artificial snare were also used by RESEARCH ON VERTEBRATES Miler and Kuszewska (2017) to check if res- cue behavior of workers of F. cinerea may be Tests consisting of a confrontation of released in response to secretions of man- two individuals were also used in the re- dibular glands, known to act as alarm pher- search on social and pro-social behavior omones in many ants (Wilson 1958; Brad- of rodents. In the present paper we pro- shaw et al. 1975; Ali et al. 1990). However, vide only a few examples of such studies. their experiments brought about negative re- Thus, D’Amato and Pavone (1993, 1996) sults. Rescue behavior directed by workers of and D’Amato (1998) showed that dyadic F. cinerea towards nestmates with impaired encounters between sibling male mice tak- mandibular gland communication (treated ing place after a relatively long separation with paint over the mandibles) did not differ (about 2 months) induce the release of opi- from responses to untreated individuals or oids that may act as a proximate reward to sham-treated ants with paint applied on mechanism for kin selection. This effect the thorax. Moreover, the tested workers did manifests itself as opioid-dependent in- not engage in rescue behavior in response crease in pain threshold that can be an- to dummy ants coated with the contents of tagonized by administration of naloxone, an crushed mandibular glands. Interestingly, two antagonist of opioid receptors. other studies devoted to the role of mandib- Another test consisting of a confronta- ular gland secretions in the elicitation of ant tion of two individuals, the so called Social rescue behavior brought about different re- Proximity test, was used in the research in- sults. Mandibular gland secretions proved to vestigating the behavior of the BTBR T+tf/J be involved in the elicitation of nestmate res- (BTBR) mice, an inbred mouse strain with cue behavior in the termite-hunting ant M. low sociability phenotype resembling the analis (Frank et al. 2017), and in the har- first diagnostic symptoms of autism:- defi vester ant V. pergandeyi responding to nest- cits in reciprocal social interactions (De- mates ensnared in spider silk (Kwapich and fensor et al. 2011; Meyza et al. 2015). As Hölldobler 2019). shown by these experiments, BTBR mice In yet another study Miler et al. avoid reciprocal frontal orientations, among (2017a) used both types of dyadic nestmate others by crawling under another mouse. rescue tests (with antlion larvae and with That behavior resembles gaze aversion, a artificial snares) to investigate the impact fundamental predictor of autism. Further of ecological niche on presence/absence of research confirmed that unavoidable social rescue behavior in behavioral repertories interactions are highly aversive for BTBR of six ant species from both tropical and mice: exposure to social proximity induces temperate regions (Camponotus korthalsiae, strong activation of periaqueductal brain Anoplolepis gracilipes and Iridomyrmex an- regions involved in the mediation of defen- ceps from Borneo, and Myrmica ruginodis, sive behavior. F. polyctena and F. cinerea from Poland). Dyadic encounters of a freely moving Rescue behavior proved to be present in potential rescuer with an entrapped vic- behavioral repertories of sand-dwelling ants tim were also used in the research inves- exposed to the risk of being captured by tigating rescue behavior and empathy in antlion larvae and of being trapped in a rodents, rats (Bartal et al. 2011, 2014, collapsed chamber of their ground nest (F. 2016; Sato et al. 2015; Blystad et al. cinerea), and ants inhabiting forest ground- 2019; Carvalheiro et al. 2019; Yamagishi cover that are also frequently exposed to et al. 2019) and mice (Ueno et al. 2019). various types of entrapment (I. anceps and During the test a freely moving individual F. polyctena). However, no rescue attempts had to liberate another individual trapped were observed in ants from species associ- in a restrainer (Bartal et al. 2011, 2014, ated with open plains, nesting in hardened 2016; Blystad et al. 2019; Carvalheiro et soils and foraging largely on herbaceous al. 2019), and in some cases also addition- plants (A. gracilipes and M. ruginodis), nor ally water-soaked (Sato et al. 2015; Yamag- in ants living in close mutualistic relation- ishi et al. 2019). ship with their host plant (C. korthalsiae). 570 Ewa Joanna Godzińska i współaut.

CONCLUSIONS Beye M., Neumann P., Chapuisat M., Pamilo P., Moritz R. F. A., 1998. Nestmate recognition and the genetic relatedness of nests in the We hope that this review demonstrated ant Formica pratensis. Behav. Ecol. Sociobiol. that even relatively simple behavioral bioas- 43, 67-72. says such as dyadic nestmate reunion tests Blystad M. H., Andersen D., Johansen E. B., may bring about interesting data. Dyadic 2019. Female rats release a trapped cagemate nestmate reunion tests applied to the study following shaping of the door opening respon- se: opening latency when the restrainer was of ant social behavior yielded and continue baited with food, was empty, or contained a to yield a wealth of important findings -ex cagemate. PloS One 14, e0223039. tending our knowledge about behavioral ef- Bonavita-Cougourdan A., 1983 Activité antenna- fects of social isolation, causal factors in- ire et flux trophallactique chez la fourmi- Cam ponotus vagus Scop. (, Formici- volved in the mediation of various social dae). Insect. Soc. 30, 423-442. contacts, nestmate recognition, role of bio- Bonavita-Cougourdan A., Gavioli M., 1981. Les genic amines in the mediation of ant social inversions du sens du flux alimentaire au co- behavior, behavioral polymorphism and divi- urs d’un meme contact entre deux ouvriéres sion of labor encountered in ant societies, chez la fourmi Camponotus vagus Scop (Hy- menoptera, Formicidae). Insect. Soc. 28, 321- ant behavioral ontogeny, diversity and vari- 340. ability of ant behavior, role of ecological fac- Bonavita-Cougourdan A., Morel L., 1984a. Les tors in the evolution of ant behavior, and activités antennaires au cours des contacts biological roots of aggressive and pro-social trophallactiques chez la fourmi Camponotus vagus Scop. ont-elles valeur de signal? Insect. behavior. Soc. 31, 113-131. Summary Bonavita-Cougourdan A., Morel L., 1984b. Poly- ethisme et comportements de relation chez les The main aim of this review was to show that even fourmis. Actes Coll. Ins. Soc. 1, 27-30. relatively simple behavioral bioassays may bring about Bonavita-Cougourdan A., Morel L., 1985. Poly- important findings. Dyadic nestmate reunion test used ethism and social interactions in ants. Behav. in the research on social insect behavior consists of a Process. 11, 425-433. confrontation of two nestmates and may be preceded by Bonavita-Cougourdan A., Morel L., 1986. Rela- various treatments such as social isolation or admin- tions entre trophallaxie, lésions des antennes et facteurs saisonniers chez la fourmi Campo- istration of neuroactive compounds. Dyadic nestmate notus vagus Scop. Insect. Soc. 33, 132-141. rescue test consists of a confrontation between a freely Bonavita-Cougourdan A., Morel L., 1988. Inter- moving ant and its endangered nestmate. Dyadic nest- -individual variability and idiosyncrasy in so- mate reunion tests continue to yield a wealth of find- cial behaviours in the ant Camponotus vagus ings extending our knowledge on behavioral effects of Scop. Ethology 77, 58-66. social isolation, causal factors involved in the mediation Bonavita-Cougourdan A., Le Masne G., Rage P., of social contacts, nestmate recognition, role of biogenic 1979. Une nouvelle methode pour l’étude des amines in the mediation of ant social behavior, behav- échanges alimentaires dans les societés de fo- ioral polymorphism and division of labor in ant socie- urmis. Insect. Soc. 26, 5-12. Bos N., Lefèvre T., Jensen A. B., D’Ettorre P., ties, ant behavioral ontogeny, diversity and variability of 2011. Sick ants become unsociable. J. Evol. ant behavior, role of ecological factors in the evolution Biol. 25, 342-351. of ant behavior, and biological roots of aggressive and Boulay R., Lenoir A., 2001. Social isolation of pro-social behavior. mature workers affects nestmate recognition in the ant Camponotus fellah. Behav. Process. 55, 67-73. REFERENCES Boulay R., Auger J., Godzińska E. J., Lenoir A., 1999a. Influence de l’octopamine sur le- com Ali M. F., Morgan E. D., 1990. Chemical com- portement trophallactique de la fourmi Campo- munication in insect communities: a guide to notus fellah (Hymenoptera, Formicidae). Actes insect pheromones with special emphasis on Coll. Ins. Soc. 12, 127-130. social insects. Biol. Rev. 65, 227-247. Boulay R., Quagebeur M., Godzińska E. J., Le- Bartal I. B. A., Decety J., Mason P., 2011. Em- noir A., 1999b. Social isolation in ants: evi- pathy and pro-social behavior in rats. Science dence of its impact on survivorship and beha- 334, 1427-1430. vior in Camponotus fellah (Hymenoptera, For- Bartal I. B. A., Rodgers D. A., Sol Bernardez micidae). Sociobiology 33, 11-134. Sarria M., Decety J., Mason P., 2014. Pro- Boulay R., Hefetz A., Soroker V., Lenoir A., -social behavior in rats is modulated by social 2000a. Camponotus fellah colony integration: experience. eLife 3, e01385. worker individuality necessitates frequent hy- Bartal I. B. A., Shan H., Molasky N. M., Mur- drocarbon exchanges. Anim. Behav. 59, 1127- ray T. M., Williams J. Z., Decety J., Mason 1133. P., 2016. Anxiolytic treatment impairs helping Boulay R., Soroker V., Godzińska E. J., Hefetz behavior in rats. Front. Psychol. 7, 850. A., Lenoir A., 2000b. Octopamine reverses the Bernadou A., Busch J., Heinze J., 2015. Diversi- isolation-induced increase in trophallaxis in ty in identity: behavioral flexibility, dominance, the carpenter ant Camponotus fellah. J. Exp. and age polyethism in a clonal ant. Behav. Biol. 203, 513-520. Ecol. Sociobiol. 69,1365-1375. Boulay R., Katzav-Gozansky T., Vander Meer R. Beye M., Neumann P., Moritz R., 1997. Nestma- K., Hefetz A., 2003. Colony insularity thro- te recognition and the genetic gestalt in the ugh queen control on worker social motivation mound-building ant Formica polyctena. Insect. in ants. Proc. Biol. Sci. 270, 971-977. Soc. 44, 49-58. Dyadic nestmate reunion test in the research on ant social behavior 571

Boulay R., Katzav-Gozansky T., Hefetz A., Leno- Duhoo T., Durand J.-L., Hollis K. L., Nowbaha- ir A., 2004. Odour convergence and tolerance ri E., 2017. Organization of rescue behaviour between nestmates through trophallaxis and sequences in ants, Cataglyphis cursor, reflects grooming in the ant Camponotus fellah (Dalla goal-directedness, plasticity and memory. Be- Torre). Insect. Soc. 51, 55-61. hav. Process. 139, 12-18. Bradshaw J. W. S., Baker R., Howse P. E., Dussutour A., Poissonnier L.-A., Buhl J., Simp- 1975. Multicomponent alarm pheromones of son S. J., 2016. Resistance to nutritional the weaver ant. Nature 258, 230-231. stress in ants: when being fat is advantage- Carvalheiro J., Seara-Cardoso A., Mesquita A. ous. J. Exp. Biol. 219, 824-833. R., de Sousa L., Oliveira P., Summavielle T., Ehrhardt S., 1931. Űber Arbeitsteilung bei Myr- Magalhães A., 2019. Helping behavior in rats mica und Messor-arten. Z. Morphol. Ökol Tie- (Rattus norvegicus) when an escape alterna- re 20, 755-812. tive is present. J. Comp. Psychol. 133, 452- Erber J., Kloppenburg P., Scheidler A., 1993. 462. Neuromodulation by serotonin and octopami- Chmurzyński J. A., 1973. Etologia a zoopsycho- ne in the honeybee: behaviour, neuroanatomy logia. [In:] Powstawanie nowych dyscyplin and electrophysiology. Experientia 49, 1073- naukowych. Geblewicz E. (ed.), Ossolineum, 1083. Wrocław-Gdańsk, 17-67. Farooqui T., Robinson K., Vaessin H., Smith B. Cichoń M., 2017. Wpływ rewersji behawioralnej H., 2003. Modulation of early olfactory proces- na zachowania ratunkowe mrówki ćmawej sing by an octopaminergic reinforcement path- (Formica polyctena Först.). M. Sc. Thesis, Fa- way in the honeybee. J. Neurosci. 23, 5370- culty of Animal Science, Warsaw University 5380. of Life Sciences WULS-SGGW. Frank E. T., Schmitt T., Hovestadt T., Mitesser Cybulska A., Godzińska E. J., 1999. Trofalaksja: O., Stiegler J., Linsenmair K. E., 2017. Sa- stary termin, nowe pytania. Wszechświat 100, ving the injured: rescue behavior in the termi- 4-6. te-hunting ant Megaponera analis. Sci. Adv. 3, Cybulska A., Godzińska E. J., Wagner-Ziemka e1602187. A., 2000. Behaviour of dyads of ants reunited Frank E. T., Wehrhahn M., Linsenmair K. E., after social deprivation. Biol. Bull. Poznań 37, 2018. Wound treatment and selective help in 119-127. a termite-hunting ant. Proc. R. Soc. B 285, Czechowski W., Godzińska E. J., Kozłowski M. 2017-2457. W., 2002. Rescue behavior shown by workers Giurfa M., Sandoz J. C., 2012. Invertebrate lear- of Formica sanguinea Latr., F. fusca L. and ning and memory: fifty years of olfactory- con F. cinerea Mayr (Hymenoptera: Formicidae) in ditioning of the proboscis extension response response to their nestmates caught by an ant in honeybees. Learn. Memory 19, 54-66. lion larva. Annal. Zool. 52, 423-431. Godzińska E. J., 1996a. Etologia owadów spo- Czechowski W., Radchenko A., Czechowska W., łecznych: fakty i kontrowersje. Kosmos 45, Vepsäläinen K., 2012. The ants of Poland 163-178. with reference to the myrmecofauna of . Godzińska E. J. 1996b. Etologia i mechanizmy Fauna Poloniae n.s. 4. Natura Optima Dux zachowania. Kosmos 45, 253-271. Foundation, Warszawa. Godzińska E. J., 1997. Etologiczna analiza za- Dahbi A., Lenoir A., 1998. Nest separation and chowania się i jego uwarunkowań przyczyno- the dynamics of the gestalt odor in the poly- wych. Kosmos 46, 181-192. domous ant Cataglyphis iberica (Hymenoptera, Godzińska E. J., 2005. Ukwiały, mrówki i ludzie: Formicidae). Behav. Ecol. Sociobiol. 42, 349- biologiczne korzenie agresji i przemocy. [In:] 355. Psychospołeczne i neurofizjologiczne aspekty D’Amato F. R., 1998. Kin interaction enhances agresji. Zagrodzka J., Kowaleczko-Szumow- morphine analgesia in male mice. Behav. ska M. (eds.). Wydawnictwo Instytutu Psycho- Pharmacol. 9, 369-373. logii PAN, Warszawa, 131-147. D’Amato F. R., Pavone F., 1993. Endogenous Godzińska E. J., 2006. Definicja i cele poznawcze opioids: a proximate reward mechanism for neurosocjobiologii. Kosmos 55, 137-148. kin selection? Behav. Neural. Biol. 60, 79-83. Godzińska E. J., 2007. Owady społeczne: mity i D’Amato F. R., Pavone F., 1996. Reunion of se- fakty. Kosmos 56, 371-381. parated sibling mice: neurobiological and be- Godzińska E. J., 2016. Human and ant social havioral aspects. Neurobiol. Learn. Mem. 65, behavior should be compared in a very care- 9-16. ful way to draw valid parallels. Behav. Brain Defensor E. B., Pearson B. L., Pobbe R. L. H., Sci. 39, e98. Bolivar V. J., Blanchard D. C., Blanchard Godzińska E. J., 2019. Etologia i co dalej? Nie- R. J., 2011. A novel social proximity test sug- które filozoficzne kwestie kształtujące badania gests patterns of social avoidance and gaze zachowania zwierząt. Filozofia i Nauka 7, 69- aversion-like behavior in BTBR T+ tf/J mice. 89. Behav. Brain Res. 217, 302-308. Godzińska E. J., Szczuka A., Korczyńska J., Delage B., 1968. Recherches sur les fourmis mo- 1999. Maximum longevity of workers of three issoneuses du Bassin Aquitain. Ethologie, phy- ant species under laboratory conditions (Hy- siologie de l’alimentation. Ann. Sci. Nat. 12e menoptera: Formicidae). Pol. Pismo Entomol. série, 10, 197-265. 38, 47-55. Delage B., Jaisson P., 1969. Étude des relations Gowdy L., Krall L., 2016. The economic origins sociales chez des fourmis du genre Aphaeno- of ultrasociality. Behav. Brain Sci. 39, e92. gaster. C. R. Acad. Sci. Paris, 268, 701-703. Hammer M., Menzel R., 1998. Multiple sites of Diez L., Urbain L., Lejeune P., Detrain C., 2015. associative odor learning as revealed by local Emergency measures: adaptive response to brain microinjections of octopamine in honey- pathogen intrusion in the ant nest. Behav. . Learn. Memory 5, 146-156. Process. 116, 80-86. Heinze J., Walter B., 2010. Moribund ants le- Dobrzańska J.,1959. Studies on the division of la- ave their nests to die in social isolation. Curr. bour in ants genus Formica. Acta Biol. Exp. Biol. 20, 249-252. 19, 57-81. 572 Ewa Joanna Godzińska i współaut.

Heinze J., Foitzik S., Hippert A., Hölldobler B., Kostowski W., 1994. Zachowanie agresywne 1996. Apparent dear enemy phenomenon and mrówek a substancje neuroprzekaźnikowe. environment-based recognition cues in the ant Wszechświat 95, 115-119. Leptothorax nylanderi. Ethology 102, 510-522. Kuraszkiewicz K., 2017. Wpływ treningu oraz Hollis K. L., Nowbahari E., 2013a. Toward a be- udział indywidualnych taktyk alternatywnych havioral ecology of rescue behavior. Evol. Psy- w kształtowaniu zachowań ratunkowych ro- chol. 11, 647-664. botnic mrówki ćmawej (Formica polyctena Hollis K. L., Nowbahari E., 2013b. A compara- Först.). M. Sc. Thesis, Faculty of Biology and tive analysis of precision rescue behaviour in Environmental Sciences, Cardinal Stefan Wy- sand-dwelling ants. Anim. Behav. 85, 537- szyński University in Warsaw. 544. Kwapich C. L., Hölldobler B., 2019. Destruc- Hölldobler B., Wilson E. O., 1990. The ants. tion of spiderwebs and rescue of ensnared Harvard University Press, Cambridge, MA. nestmates by a granivorous desert ant (Ve- Hölldobler B., Wilson E. O., 1998. Podróż w romessor pergandei). Am. Nat. 194, doi: krainę mrówek. Dzieje badań naukowych. Wy- 10.1086/704338. dawnictwo Prószyński i Spółka, Warszawa. Leclerc J.-B., Detrain C., 2017. Loss of attrac- Hölldobler B., Wilson E. O., 2009. The superor- tion for social cues leads to fungal-infected ganism. The beauty, elegance and strangeness Myrmica rubra ants withdrawing from the of insect societies. W. W. Norton, New York, nest. Anim. Behav. 129, 133-141. London. Le Moli F., Mori A., 1986. The aggression tests Huang Z.-Y., Robinson G. E., 1996. Regulation as a possible taxonomic tool in the Formica of honey bee division of labor by colony age rufa group. Aggressive Behav. 12, 93-102. demography. Behav. Ecol. Sociobiol. 39, 147– Le Moli F., Parmigiani S., 1982. Intraspecific 158. combat in the red wood ant (Formica lugubris Ichinose K., Cerdá X., Christidès J.- P., Lenoir Zett.). Aggressive Behav. 8, 145-148. A., 2005. Detecting nestmate recognition pat- Le Moli F., Mori A., Parmigiani S., 1982. Agoni- terns in the fission-performing ant Aphaenoga- stic behaviour of L. (Hymenopte- ster senilis: a comparison of different indices. ra Formicidae). Monit. Zool. Ital. 16, 325-331. J. Insect. Behav. 18, 633-650. Le Moli F., Mori A., Parmigiani S., 1983a. In- Jaisson P., 1993. La fourmi et le sociobiologiste. terspecific aggression between Formica rufa Editions Odile Jacob, Paris. L. and Formica cunicularia Latr. (Hymenopte- Jemielity S., Chapuisat M., Parker J. D., Keller ra Formicidae). Atti. Soc. Ital. Sci. Nat. Mus. L., 2005. Long live the queen: studying aging Civ. Stor. Nat. Milano 124, 29-37. in social insects. Age 27, 241-248. Le Moli F., Mori A., Parmigiani S.,1983b. The ef- Jutsum A. R., Saunders T. S., Cherrett J. M., fect of antennalectomy on attack behaviour of 1979. Intraspecific aggression in the leaf-cut- Formica lugubris Zett. (Hymenoptera: Formici- ting ant Acromyrmex octospinosus. Anim. Be- dae). Ital. J. Zool. 50, 201-206. hav. 27, 839-844. Le Moli F., Mori A., Parmigiani S., 1984. Studies Katzav-Gozansky T., Boulay R., Vander Meer R., on interspecific aggression among red wood Hefetz A., 2004. In-nest environment modula- ant species Formica rufa L. vs Formica lugu- tes nestmate recognition in the ant Campono- bris Zett. (Hymenoptera Formicidae). Monit. tus fellah. Naturwissenschaften 91, 186-90. Zool. Ital. 18, 41-51. Keller L., 1998. Queen lifespan and colony cha- Lenoir A., 1979a. Le comportement alimentaire et racteristics in ants and termites. Insect. Soc. la division du travail chez la fourmi Lasius ni- 45, 235-246. ger. Bull. Biol. Fr. Belg. 113, 79-314. Keller L., Genoud M., 1997. Extraordinary life- Lenoir A., 1979b. Feeding behaviour in young so- spans in ants: a test of evolutionary theories cieties of the ant Tapinoma erraticum L.: tro- of ageing. Nature 389, 958-960. phallaxis and polyethism. Insect. Soc. 26, 19- Kleineidam C. J., Heeb E. L., Neupert S., 2017. 37. Social interactions promote adaptive resource Lenoir A., 1982. An informational analysis of an- defense in ants. PloS One 12, e0183872. tennal communication during trophallaxis in Korczyńska J., 1996. Rola czynników nabytych the ant Myrmica rubra L. Behav. Process. 7, w ontogenezie preferencji środowiskowych i 27-35. pokarmowych mrówek. Kosmos 45, 419-432. Lenoir A., Cuisset B., Hefetz A., 2001a. Effects Korczyńska J., 2001. Czynniki wpływające na of social isolation on hydrocarbon patterns preferencje warunków oświetlenia u mrówek. and nestmate recognition in the ant Aphaeno- Ph. D. Thesis. Nencki Institute of Experimen- gaster senilis (Hymenoptera, Formicidae). In- tal Biology of the Polish Academy of Scien- sect. Soc. 48, 101-109. ces, Warsaw. Lenoir A., Hefetz A., Simon T., Soroker V., Korczyńska J., Szczuka A., Kieruzel M., Maj- 2001b. Comparative dynamics of gestalt odour czynski H., Khvorostova N., Godzińska E. J., formation in two ant species Camponotus fel- 2005. Effects of the biogenic amines, dopami- lah and Aphaenogaster senilis (Hymenoptera: ne, tyramine and octopamine on the behavior Formicidae). Physiol. Entomol. 26, 275-283. of carpenter ant workers [Camponotus hercu- Leonhardt S. D., Menzel F., Nehring V., Schmitt leanus (Hymenoptera: Formicidae)] during nest- T., 2016. Ecology and evolution of communica- mate reunion tests carried out after a period tion in social insects. Cell 164, 1277-1287. of social isolation. Sociobiology 45, 409-447. Mazurkiewicz P. J., Wagner-Ziemka A., Mirecka Korczyńska J., Szczuka A., Symonowicz B., Wnuk A., Godzińska E. J., 2016. Behaviour of in- A., Gonzalez Szwacka A., Mazurkiewicz P., tranidal and extranidal major workers of the Studnicki M., Godzińska E. J., 2014. The ef- African carpenter ant Camponotus maculatus fects of age and past and present behavioral Fabricius (Hymenoptera: Formicidae) during specialization on behavior of workers of the dyadic nestmate reunion tests. Afr. Entomol. red wood ant Formica polyctena Först. during 24, 307-320. nestmate reunion tests. Behav. Process. 107, Mazurkiewicz P. J., Szczepanik J., Wild A., Nie- 24-41. lubowicz M., Godzińska E. J., 2015. Mrówka Dyadic nestmate reunion test in the research on ant social behavior 573

na każdy dzień. Wydawnictwo SCRIPT, War- Roeder T., 2005. Tyramine and octopamine: ru- szawa. ling behavior and metabolism. Annu. Rev. En- McDonald P., Topoff H., 1985. Social regulation tomol. 50, 447-477. of behavioral development in the ant, Novo- Roeder T., Seifert M., Kähler C., Gewecke M., messor albisetosus (Mayr). J. Comp. Psychol. 2003. Tyramine and octopamine: antagonistic 99, 3-14. modulators of behavior and metabolism. Arch. Mercier J.-L., Lenoir A., Déjean A., 1997. Ritu- Ins. Biochem. Physiol. 54, 1-13. alised versus aggressive behaviours display- Sato N., Tan L., Tate K., Okada M., 2015. Rats ed by Polyrhachis laboriosa (F. Smith) during demonstrate helping behavior toward a so- intraspecific competition. Behav. Process. 41, aked conspecific. Anim. Cogn. 18, 1039-1047. 39-50. Shpigler H. Y., Saul M. C., Corona F., Block Meyza K., Nikolaev T., Kondrakiewicz K., Blan- L., Ahmed A. C., Zhao S. D., Robinson G.E., chard D. C., Blanchard R. J., Knapska E., 2017. Deep evolutionary conservation of au- 2015. Neuronal correlates of asocial behavior tism-related genes. Proc. Natl. Acad. Sci. USA in a BTBR T+Itpr3tf /J mouse model of au- 114, 9653-9658. tism. Front. Behav. Neurosci. 9,199. Sorensen A. A., Busch T. M., Vinson S. B., Miler K., 2016. Moribund ants do not call for 1984. Behavioral flexibility of temporal subca- help. PloS One 11, e0151925. stes in the fire ant, Solenopsis invicta -in re Miler K., Kuszewska K., 2017. Secretions of sponse to food. Psyche 91, 319-331. mandibular glands are not involved in the eli- Symonowicz B., Kieruzel M., Szczuka A., Kor- citation of rescue behaviour in Formica cinerea czyńska J., Wnuk A., Mazurkiewicz P. J., ants. Insect. Soc. 64, 303-305. Chiliński M., Godzińska E. J., 2015. Behavio- Miler K., Bakhtiar E.Y., Czarnolesi M., 2017a. ral reversion and dark-light choice behavior in Pro-social behaviour of ants depends on the- workers of the red wood ant Formica polycte- ir ecological niche - Rescue actions in species na. J. Insect Behav. 28, 245-256. from tropical and temperate regions. Behav. Szczuka A., 1996. Elastyczność behawioralna Process. 144, 1-4. mrówek. Kosmos 45, 433-442. Miler K., Symonowicz B., Godzińska E. J., Szczuka A., Godzińska E. J., 1997. The effect of 2017b. Increased risk proneness or social wi- past and present group size on responses to thdrawal? The effects of shortened life expec- prey in the ant Formica polyctena Först. Acta tancy on the expression of rescue behavior in Neurobiol. Exp. 57, 135-150. workers of the ant Formica cinerea (Hymenop- Szczuka A., Godzińska E. J., 2000. Group size: tera: Formicidae). J. Insect Behav. 30, 632- an important factor controlling the expression 644. of predatory behaviour in workers of the wood Moffett M. W., 2018. The human swarm. How ant Formica polyctena Först. Biol. Bull. Po- our societies arise, thrive, and fall. Basic Bo- znań 37, 139-152. oks, New York. Szczuka A., Godzińska E. J., 2004a. The role of Morel L., 1986. Comportement trophallactique de group size in the control of expression of pre- l’ouvrière agée face á une immature chez la datory behavior in workers of the red wood fourmi Camponotus vagus Scop. Insect. Soc. ant Formica polyctena (Hymenoptera: Formici- 33, 32-44. dae). Sociobiology 43, 295-325. Moroń D., Witek M., Woyciechowski M., 2008. Szczuka A., Godzińska E. J., 2004b. The effect of Division of labour among workers with diffe- gradual increase of group size on the expres- rent life expectancy in the ant Myrmica scabri- sion of predatory behavior in workers of the nodis. Anim. Behav. 75, 345-350. red wood ant Formica polyctena (Hymenopte- Moroń D., Lenda M., Skórka P., Woyciechowski ra: Formicidae). Sociobiology 43, 327-349. M., 2012. Short-lived ants take greater risks Szczuka A., Godzińska E. J., 2008. Effects of during food collection. Am. Nat. 180,744-750. chronic oral administration of octopamine on Nowbahari E., Hollis K. L., 2010. Rescue be- the expression of predatory behavior in small havior: distinguishing between rescue, coope- groups of workers of the red wood ant For- ration, and other forms of altruistic behavior. mica polyctena (Hymenoptera: Formicidae). So- Commun. Integr. Biol. 3, 1-3. ciobiology 52, 703-728. Nowbahari E., Amirault C., Hollis K . L., 2016. Szczuka A., Korczyńska J., Wnuk A., Symono- Rescue of newborn ants by older Cataglyphis wicz B., Gonzalez Szwacka A., Mazurkiewicz cursor adult workers. Anim. Cogn. 19, 543- P., Kostowski W., Godzińska E. J., 2013. 553. The effects of serotonin, dopamine, octopamine Nowbahari E., Hollis K. L., Durand J.-L., 2012. and tyramine on behavior of workers of the Division of labor regulates precision rescue ant Formica polyctena during dyadic aggres- behavior in sand dwelling Cataglyphis cursor sion tests. Acta Neurobiol. Exp. 73, 495-520. ants. PloS One 7, e48516. Taylor K., Visvader A., Nowbahari E., Hollis K. Nowbahari E., Scohier A., Durand J.-L., Hol- L., 2013. Precision rescue behavior in North lis K. L., 2009. Ants, Cataglyphis cursor, use American ants. Evol. Psychol. 11, 665-677. precisely directed rescue behavior to free en- Thorpe W. H., 1979. The origins and rise of etho- trapped relatives. PloS One 4, 65-73. logy. Heinneman, London. Parmigiani S., Le Moli F., 1987. Relationships Tinbergen N., 1963. On aims and methods of between intra and intrspecific aggression: the ethology. Z. Tierpsychol. 20, 410-433. case of Formica rufa group species. Pubblica- Ueno H., Suemitsu S., Murakami S., Kitamura N., zioni dell’ Instituto di Entomologia dell’Uni- Wani K., Matsumoto Y., Okamoto M., Ishi- versita di Pavia, 36, 29-32. hara T., 2019. Helping-like behaviour in mice Pribbenow B., Erber J., 1996. Modulation of an- towards conspecifcs constrained inside tubes. tennal scanning in the honeybee by sucrose Sci. Rep. 9, 10648. stimuli, serotonin, and octopamine: behavior Ugelvig L. V, Cremer S., 2007. Social prophy- and electrophysiology. Neurobiol. Learn. Me- laxis: group interaction promotes collective im- mory 66, 109-120. munity in ant colonies. Curr. Biol. 17, 1967- 1971. 574 Ewa Joanna Godzińska i współaut.

Uy F. M. K., Adcock J. D., Jeffries S. F., Pe- Wilson E. O., 1958. A chemical releaser of alarm pere E., 2019. Intercolony distance predicts and digging behavior in the ant Pogonomyr- the decision to rescue or attack conspecifics in mex badius. Psyche 65, 41-51. weaver ants. Insect. Soc. 66, 185-192. Wilson E. O., 1971. The insect societies. Belk- Vander Meer R. K., Preston C. A., Hefetz A., nap/Harvard University Press, Cambridge, 2008. Queen regulates biogenic amine level MA. and nestmate recognition in workers of the Wilson E. O., 1975. Sociobiology. The new syn- fire ant, Solenopsis invicta. Naturwissenscha- thesis. Belknap, Cambridge, MA. ften 95, 1155-1158. Wilson E. O., Eisner T., 1957. Quantitative stu- Wada-Katsumata A., Yamaoka R., Aonuma H., dies of liquid food transmission in ants. Insect 2011. Social interactions influence dopamine Soc. 4, 157-166. and octopamine homeostasis in the brain of Wnuk A., 2013. Neurobiologiczne korelaty rozwoju the ant Formica japonica. J. Exp. Biol. 214, behawioralnego robotnic owadów społecznych. 1707-1715. Kosmos 62, 513-523. Wagner-Ziemka A., Szczuka A., Korczyńska J., Wnuk A., Godzińska E. J., 2006. Wpływ izolacji Kieruzel M., Godzińska E. J., 2006. Behavior społecznej na fizjologię i zachowanie się- mró of ant-workers of Aphaenogaster senilis (Hy- wek. Kosmos 55, 177-196. menoptera: Formicidae) during dyadic nestma- Wnuk A., Wiater M., Godzińska E. J., 2011. Ef- te reunion tests carried out after a period of fect of past and present behavioural specia- social isolation. Sociobiology 48, 281-308. lization on brain levels of biogenic amines in Wagner-Ziemka A., Gonzalez Szwacka A., Kor- workers of the red wood ant Formica polycte- czyńska J., Kieruzel M., Fiałkowska B., Go- na. Physiol. Entomol. 36, 54-61. dzińska E. J., 2008. Comparison of the beha- Yamagishi A., Okada M., Masuda M., Sato N., vior of nurses and foragers of the carpenter 2019. Oxytocin administration modulates ant, Camponotus melanocnemis, during dyadic rats’ helping behavior depending on social nestmate reunion tests carried out after a pe- context. Neurosci Res, doi: 10.1016/j.neu- riod of social isolation (Hymenoptera: Formici- res.2019.04.001. dae). Sociobiology 52, 667-700.

KOSMOS Vol. 68, 4, 561–574, 2019

Ewa Joanna Godzińska, Julita Korczyńska, Anna Szczuka

Pracownia Etologii, Instytut Biologii Doświadczalnej im. M. Nenckiego PAN, Pasteura 3, 02-093 Warszawa E-mail: [email protected]

TEST SPOTKANIA DWÓJKI TOWARZYSZEK W BADANIACH ZACHOWAŃ SPOŁECZNYCH MRÓWEK

Streszczenie Głównym celem tego przeglądu było ukazanie, że nawet stosunkowo proste testy behawioralne mogą umożliwić uzyskanie ważnych wyników. Test spotkania dwójki towarzyszek stosowany w badaniach zachowań społecznych mró- wek polega na konfrontacji dwóch towarzyszek z tej samej kolonii i może być poprzedzany różnymi zabiegami, taki- mi jak izolacja społeczna lub podawanie związków neuroaktywnych. Dwójkowy test badający zachowania ratunkowe polega na konfrontacji swobodnie poruszającej się mrówki z towarzyszką znajdującą się w niebezpieczeństwie. Testy spotkania dwójki towarzyszek umożliwiają uzyskiwanie ogromnego bogactwa wyników poszerzających naszą wiedzę o behawioralnych efektach izolacji społecznej, uwarunkowaniach przyczynowych różnych typów kontaktów społecznych, rozpoznawaniu towarzyszek z tej samej kolonii, roli amin biogennych w sterowaniu zachowaniami społecznymi mró- wek, polimorfizmie behawioralnym i podziale pracy w społeczeństwach mrówek, ontogenezie behawioralnej mrówek, różnorodności i zmienności zachowań mrówek, roli czynników ekologicznych w ewolucji zachowań mrówek, oraz bio- logicznych korzeniach zachowań agresywnych i prospołecznych.

Słowa kluczowe: behawior, Formicidae, mrówki, test spotkania dwójki towarzyszek, zachowania społeczne