Elsevier

Physiology & Behavior

Volume 179, 1 October 2017, Pages 220-225
Physiology & Behavior

Mammalian nonapeptides activate territorial behavior in an amphibian

https://doi.org/10.1016/j.physbeh.2017.06.020Get rights and content

Highlights

  • VT activates calling and territorial behavior in anuran amphibians (frogs).

  • Exogenous application of VP and OT in frogs activates calling and territoriality.

  • In silent, non-calling adult male frogs VT, OT, and VP activate territorial calling followed by movement into new territories.

Abstract

Arginine vasopressin (VP) and oxytocin (OT) are two nonapeptides present in mammals and execute a wide array of physiological and behavioral functions. In amphibians arginine vasotocin (VT) is hypothesized as a homologous nonapeptide for VP and also performs physiological and behavioral tasks. Studies have demonstrated that the structural and functional relationships between VP, OT, and VT receptor families are similar; however, little behavioral data has complimented these studies. The objective of this investigation was to determine if the mammalian nonapeptides VP and OT would activate behavioral manifestations naturally activated by VT. Frogs are particularly attractive for such an investigation because it is well documented that VT activates advertisement calling and territorial behavior. This investigation was a large sample size field study that utilized the territorial frog, Eleutherodactylus coqui. Fieldwork occurred on the Islands of Puerto Rico and Hawai'i and focused on territorial (calling) and non-territorial (silent) males. Frogs were administered exogenous injections of VP, OT, VT (positive control), or saline (control) in the field, placed back in their original locations, and were observed for behaviors. Exogenous injections of VP and OT significantly activated silent males to emit advertisement calls and exhibit territorial behavior. Additionally, silent males moved into new areas prior to calling whereas territorial males remained in their own territories. Control (saline) males displayed normal behaviors. This is the first study to demonstrate that mammalian nonapeptides activate calling and territorial behaviors in frogs and corroborates the close evolutionary relationships within the nonapeptide family.

Introduction

Nonapeptides are an archaic family of conserved peptides that have evolved for > 700 million years. Functional properties of nonapeptides and their phylogenetic dispersion within vertebrate clades are wide-ranging and diverse [1], and yet the biochemical differences among the nonapeptides are remarkably minor [2]. Vasopressin- and oxytocin-related nonapeptides are present in representatives of both protostomian and deuterostomian lineages [3], [4], [5], [6], [7], [8]. This suggests that this signaling system originated very early in metazoan evolution. Due to the structural and positional similarities of the vasopressin and oxytocin genes it is hypothesized that they originated from the duplication of a common ancestral gene, likely following the radiation of the jawless fish about 500 million years ago [3], [6], [9], [10]. This genetic event is certainly plausible since gene duplication is a common evolutionary pathway toward the adaptation of genes to new functions [11].

Arginine vasopressin (VP) and oxytocin (OT) are two nonapeptides that are present in mammals and execute a wide array of physiological and behavioral functions [12]. In other classes of vertebrates these two neuropeptide systems are not present but homologous neuropeptide systems are existent. Instead of VP, arginine vasotocin (VT) occurs in birds, reptiles, amphibians, and fish [2], [3]. Co-evolving with these peptides were their respective receptors and signaling pathways that are responsible for conducting molecular and cellular functions.

Functionally, nonapeptides execute an assortment of physiological and behavioral tasks. Traditionally, VP is known as an antidiuretic hormone for its role in water retention [13], vasoconstriction [14], and water homeostasis [15]. Meanwhile OT is classically recognized for its physiological role during parturition and lactation [16], [17], [18]. Intriguingly, a large volume of literature has been generated on the behavioral outcomes of nonapeptides. While substantial variation ensues between taxa, all linages of vertebrates are characterized by having specific behaviors governed by these peptides that can include: aggression, agonistic behavior, pair-bond formation, vocalizations, gregariousness, cooperation, and paternal and/or maternal care [for reviews see: [1], [3], [19], [20], [21], [22]].

A number of investigations (and reviews) on nonapeptides have concentrated on the functional aspects of nonapeptide systems [23], [24], [25], [26], [27]. While the majority of these studies have utilized mammals as their model organism some have employed amphibians [28], [29] and fish [30], [31]. For example, it was discovered replacing the fish gene for isotocin (teleost homolog of OT) with the mammalian gene for OT in transgenic rats did not adversely affect their physiology [31]. Further, it was demonstrated that the mammalian nonapeptides, OT and VP, can modulate social behavior in fish [32]. This could indicate that receptor mechanisms and signaling factors mediating the physiological regulation of nonapeptides are possibly conserved between mammals and fish. Investigations in both newts and frogs disclosed that both mesotocin and vasotocin 1a nonapeptide receptors are present [24], [28], [29]. In amphibians, the VT system activates indispensable social and reproductive behaviors [33], [34], [35], [36], [37] and it is firmly established that in male frogs VT activates territorial and reproductive behaviors [38]. In fact, previous research in the Puerto Rican coquí frog, Eleutherodactylus coqui, has demonstrated that VT will activate advertisement vocalizations [37]. These are signals used in the establishment and maintenance of territories as well as the attraction of mates. This investigation also found that exogenously activated, non-territorial frogs moved into and established new territories and commenced advertisement calling [39].

Clearly, nonapeptides perform a key role in the expression of social behaviors and that distinctive classes of nonapeptides exist in different clades of vertebrates. The objectives of this investigation were to 1) determine if the closely related non-amphibian nonapeptides, VP and OT, would activate territorial behavior in male Puerto Rican coquí frogs, E. coqui; 2) determine if non-territorial frogs move into new territories and commence advertisement calling, and 3) to elucidate, if VP and/or OT activate additional and/or alternative social or reproductive behaviors. Due to functional and structural components of nonapeptide receptor-ligand binding properties it is hypothesized that VP and OT will activate advertisement calling and territorial behaviors in E. coqui.

Section snippets

Field site locations

Field work was carried out at two locations: 1) the Caribbean National Forest located in the Luquillo Mountains of northeastern Puerto Rico, approximately 1 km east of the El Verde Field Station (350–400 m) of the University of Puerto Rico at Rio Piedras; and 2) the Waiakea Field Research Area (500 m) of the University of Hawai‘i at Hilo, 924 Stainback Rd., Hilo, HI. Permits were obtained from the Departmento de Recursos Naturales y Ambientales of Puerto Rico and from the Department of Land and

Advertisement call activation

Activation of advertisement calling was significantly higher in VP-injected silent males (33/50) compared to saline-injected silent males (0/50) (p < 0.00001, Fisher's Exact Test) (Table 1; Fig. 1). VP-injected territorial males (42/50) were not significantly more activated to call than saline-injected territorial males (37/50) (p = 0.326, Fisher's Exact Test) but did have significantly higher call activation than saline-injected silent males (0/50) (p < 0.00001, Fisher's Exact Test). There was not a

Discussion

Results indicate that the mammalian nonapeptides VP and OT significantly activated advertisement calling and territorial behavior in the anuran amphibian, E. coqui. Further, the VP- and OT-injected males were as significantly activated to emit advertisement frogs as VT activated males. Interestingly, a major difference between exogenously activated males was that silent males relocated into a new area prior to initiating advertisement calling significantly more often than territorial males in

Conclusions

The mammalian neuropeptides VP and OT significantly activated advertisement calling and territorial behavior in male E. coqui. This activation was as robust as activation accomplished with the amphibian nonapeptide VT. Interestingly, nonapeptide activation also affected males differently. Following exogenous injections of VP and OT territorial males remained in their current territory but silent males moved into new areas and only then began to emit advertisement calls; this difference in male

Acknowledgments

We are grateful to the Departmento de Recursos Naturales y Ambientales of Puerto Rico for issuing collecting permits to work on E. coqui. Very special thanks goes to the staff at the El Verde Field Station (EVFS), Puerto Rico for providing support and use of their facilities, especially to Dr. Alonso Ramírez (Director of EVFS), Dr. Jill Thompson, and Ms. Hilda Lugo. The field station is part of the Long-Term Ecological Research project of the Institute for Tropical Ecosystem Studies, University

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