Chem. Senses 27: 91-94,
2002
© Oxford University Press 2002
SHORT REPORT |
Disruption of the Fifth Melanocortin Receptor Alters the Urinary Excretion of Aggression-modifying Pheromones in Male House Mice
Department of Biology, Georgia State University, Atlanta, GA 30303, USA 1 Department of Biology, The University of North Carolina at Greensboro, Greensboro, NC 27402, USA
Correspondence to be sent to: Dr John Lepri, The University of North Carolina at Greensboro, Department of Biology, 312 Eberhart Life Sciences Building, Greensboro, NC 27402-6174, USA. e-mail: jjlepri{at}uncg.edu
Abstract
The preputial glands of house mice express the gene for the fifth melanocortin receptor (MC5-R) and are a primary source of urinary pheromones involved in inter-male aggression. A `resident-intruder' behavioral model was used to compare the responses of resident males to urine from mice with an engineered disruption of the fifth melanocortin receptor (MC5-RKO) with residents' responses to urine from wild-type mice (WT). Each type of urine was presented in combination with a castrated intruder male to provide the appropriate biological context. Resident males responded with a longer latency to bite when the urine was from gonadally intact WT males compared with urine from MC5-RKO mice. These results are consistent with the hypothesis that activation of the fifth melanocortin receptor in the preputial glands of male house mice causes excretion of urinary pheromones that delay aggressive responses by other males.
Introduction
Inter-male aggression is a key component in the reproductive ecology of
male house mice, where it influences the partitioning of food, mates and
shelter (Anderson and Hill,
1965
; Bronson,
1979). An individual mouse's propensity to engage in aggressive
behavior is influenced by genetic background, previous fighting experience,
body mass and the perception of aggression-related odor cues (Sandnabba,
1985;
1986), including urinary
pheromones from the preputial glands.
The paired preputial glands of house mice are modified sebaceous glands
located near the base of the penis. The size of these glands is testosterone
dependent, whereas acute secretory activity appears to be regulated by
hormones from the anterior pituitary gland, specifically by
-melanocyte-stimulating hormone (-MSH)
(Thody and Shuster, 1973). It
has been demonstrated that chemical cues secreted into the urine from the
preputial glands modify aggressive interactions
(McKinney and Christian,
1970). Socially dominant male mice have larger preputial glands
than do subordinate males (Bronson and
Marsden, 1973). Moreover, preputial-glandectomized males fail to
establish social dominance as readily as sham-operated males
(Ingersoll et al.,
1986; Hayashi,
1987). These and other results have been interpreted to suggest
that the preputial glands are the source of a chemical signal that promotes
aggression. However, the current study demonstrates that robust preputial
signals might cause a recipient of these signals to delay the onset of
aggressive behavior, thus reducing the recipient's risk of physical injury
resulting from fighting with a dominant male.
The fifth melanocortin receptor (MC5-R) is a 7-transmembrane G-coupled
protein receptor that is found in a variety of exocrine glands, including the
preputial glands (Chen et al.,
1997; Tatro and Reichlin,
1987). The endogenous ligands for the MC5-R are the melanocortins,
including -MSH (Grantz et
al., 1994; Griffon et
al., 1994; Labbe et
al., 1994; Fathi et
al., 1995; Hruby et
al., 1995). The availability of mice with a disruption of the
fifth melanocortin receptor (MC5-RKO) provided a unique opportunity to examine
the pheromonal contribution of the preputial glands to urine. Because the
preputial glands are the only component of the urogenital system known to
express the MC5-R gene, it was hypothesized that its disruption would alter
the aggression-modifying qualities of urine from MC5-RKO male mice.
Materials and methods
Strains of Mus musculus domesticus (Taconic) were used in this
experiment: ICR male `retired breeders' were used as resident males
(n = 16), and castrated males of the C57/B6 strains were used as
intruders (n = 3). Gonadally intact WT (n = 6) and MC5-RKO
(n = 6) male mice of the C57/B6 strain provided stimulus urine. The
production of the knockout mice is described by Chen et al.
(Chen et al., 1997).
All mice were individually housed at the University of North Carolina at
Greensboro Animal Facility in polypropylene cages of size 25 x 20
x 15.5 cm. Animals were maintained in a 14:10 light:dark reverse light
cycle, with food (Harlan TEKLAD Rodent Diet) and tap water provided ad
libitum. All testing was conducted during the dark part of the light:dark
cycle in recognition of the nocturnal behavior patterns of house mice.
Proposals for all aspects of this work were institutionally reviewed and
approved.
A modified resident—intruder model was used to compare the aggressive responses of `resident' males to urine previously collected from WT males and MC5-RKO males. The urine had been collected, pooled by genotype and frozen at -20°C until tested. Resident males were prescreened for aggressive behavior 1 week prior to testing by dangling a mouse in front of them for up to 1 min on four consecutive days; residents were classified as aggressive if they attacked the dangling mouse on one or more of these four days. Sixteen of 20 males were deemed `aggressive' based on these criteria.
To provide the biological context for testing pheromone activity, castrated `intruder' males were placed into a urine-scented protective cage, hereafter called the `corral', which was a double-walled cylindrical chamber made of hardware cloth. Thirty microliters of urine from either WT or MC5-RKO males was pipetted onto clean paper and wire `twist ties' attached to the ends of the corral. Qualitative observations made prior to this experiment verified that resident males did not attack the corral in the absence of the intruder male or urine.
The behavioral tests were 15 min in duration and several behavioral measurements were recorded. Measurements of aggressive behavior included latencies (time elapsed to first occurrence) and frequencies (total number) of `bites' (resident's mouth placed on the wires of the corral) and `attacks' (resident's front paws placed on the corral simultaneously with a vigorous bite). To compare baseline activity in the presence of different odor stimuli, locomotor activity (number of times the resident's nose crossed a midline) was measured. Finally, in an attempt to ascertain the interaction of the residents with the urine stimuli, the following measurements were recorded: climbing onto the top of the corral, and latencies and durations of proximity (resident's nose within 1 cm) to the corral and to the stimuli. A latency score of 900 s (the duration of the test) was assigned for any behaviors that did not occur.
In random order, over 2 days, each of the 16 resident male subjects was
tested with each type of stimulus urine. The type of urine tested was unknown
to the observer. The raw behavioral responses of the residents were analyzed
by one-way analysis of variance for genotypic differences, using JMP
statistical software (SAS Institute,
1995).
Results
Urine from MC5-RKO mice was associated with a significantly decreased latency to bite [F(1,32) = 6.7940; P = 0.02; mean latencies to bite ± SEM were 319.7 ± 89.6 versus 142.5 ± 36.8 s for WT and MC5-RKO, respectively; see Figure 1]; and a significant decrease in the number of times the residents climbed on top of the corral [F(1,32) = 6.7960; P = 0.02; mean frequencies ± SEM were 23.5 ± 1.5 versus 18.6 ± 1.8 observations for WT and MC5-RKO, respectively]. There were no statistically significant differences in locomotor activity, or in the latency and duration of time spent in proximity to the stimuli, indicating that all subjects had equivalent access to the stimuli. In addition, there were no significant differences in the total number of bites or attacks, indicating that the primary effects of disrupting MC5-R were to alter the excretion of signals that modify the onset, rather than full expression, of aggressive behaviors.
|
Discussion
This experiment demonstrated that the urine from MC5-RKO mice was
associated with accelerated onset of aggressive behavior compared with urine
from WT mice. This result is consistent with earlier experiments in which
urine from `stressed' and `unstressed' MC5-RKO mice was associated with a
decreased latency to bite and attack compared with urine from WT mice
(Caldwell et al.,
2001). Over the full duration of the tests, there were no
differences in the frequency of bites and attacks in response to the urine
from the two genotypes, suggesting that the delayed onset of aggressive
behavior in the presence of WT urine does not result in an overall reduction
in the display of aggressive behavior. However, the rapid onset of aggressive
responses could influence the outcome of interactions between males.
The preputial glands are the only urogenital glands known to express MC5-R
(Tatro and Reichlin, 1987;
Labbe et al., 1994;
Chen et al., 1997), so
they are likely to be the source of apparent `aversion pheromones' or
aggression-inhibiting pheromones excreted by dominant male mice. The idea that
these signals inhibit or reduce inter-male aggression is supported by previous
experiments demonstrating that male mice avoid cage areas that have been
`scent-marked' with urine from a dominant male
(Jones and Nowell, 1973).
Specifically, the farnesenes are likely candidates for the such chemosignals
because - and ß- isomers of the farnesenes, while absent in
bladder urine, are present in the preputial glands and in the excreted urine
of dominant mice (Novotny et al.,
1990). As a behavioral demonstration of the effects of farnesenes,
it was shown that socially subordinate male mice were attracted to test areas
`marked' with urine from female mice; however, when exposed to female urine to
which farnesene has been added, mice avoided the test area
(Novotny et al.,
1990). Additional support that WT mice can excrete chemosignals
absent in MC5-RKO urine comes from a recent report that MC5-RKO mice have a
reduction in volatile compounds, including farnesenes, in their preputial
gland extracts (Morgan et al.,
1999). Direct tests of farnesene isomers using the
resident—intruder model could reveal their possible role in delaying the
onset of aggression.
In summary, these data support the hypothesis that melanocortins control the excretion of preputial-gland products that may function to delay the onset of aggression. In house mouse populations, the deployment of such aversion pheromones might allow the most aggressive males to maintain dominance in a social hierarchy, with fewer adversaries initiating high-risk aggressive encounters culminating in physical attacks.
Acknowledgments
The authors thank Roger Cone and Wenbiao Chen for providing the MC5-RKO
mice, and Marjut
Mäkelä and Dave
Ludwig for technical assistance. We also thank Cheryl Logan for comments on
the manuscript. This research was supported by the UNCG Biology Department. A
preliminary report of data was presented at the 24th Annual Meeting of the
Association for Chemoreception Sciences, Sarasota, FL, 1998. A portion of this
work was used by the senior author to satisfy the requirements for a master's
degree at UNCG (Kingsley,
1998).
References
Anderson, P.K. and Hill, J.L. (1965) Mus
musculus: experimental induction of territory formation.Science
, 148,1753
-1755.
Bronson, F.H. (1979) The reproductive ecology of the house mouse. Q. Rev. Biol.,54 , 265-299.[Medline]
Bronson, F.H. and Marsden, H.M. (1973) The preputial gland as an indicator of social dominance in male mice.Behav. Biol. , 9,625 -628.[Web of Science][Medline]
Caldwell, H.K., Wang, L. and Lepri, J.J. (2001) Simplified tests of aggression chemosignals in male house mice suggest that a melanocortin-dependent product of the preputial gland reduces attacks. In Marchlewska-Koj, A., Lepri, J.J. and Muller-Schwarze, D. (eds), Chemical Signals in Vertebrates 9. Kluwer/Academic Press, New York, pp.445 -450.
Chen, W., Kelly, M.A., Opitz-Araya, X., Thomas, R.E., Low, M.J. and Cone, R.D. (1997) Exocrine gland dysfunction in MC5-R-deficient mice: evidence for coordinated regulation of exocrine gland function by melanocortin peptides. Cell,91 , 789-798.[Web of Science][Medline]
Fathi, Z., Iben, L.G. and Parker, E.M. (1995) Cloning, expression, and tissue distribution of a fifth melanocortin receptor subtype. Neurochem. Res.,20 , 107-113.[Web of Science][Medline]
Grantz, I., Shimoto, Y., Konda, Y., Miwa, H., Dickinson, C. and Yamada, T. (1994) Molecular cloning, expression, and characterization of a fifth melanocortin receptor. Biochem. Biophys. Res. Commun., 200,1214 -1220.[Web of Science][Medline]
Griffon, N., Mignon, V., Facchinetti, P., Diaz, J., Schwartz, J. and Sokoloff, P. (1994) Molecular cloning and characterization of the rat fifth melanocortin receptor. Biochem. Biophys. Res. Commum., 200,1007 -1014.[Web of Science][Medline]
Hayashi, S. (1987) The effects of preputialectomy on aggression in male mice. Zool. Sci.,4 , 551-555.[Web of Science]
Hruby, V.J., Lu, D., Sharma, S.D., Castrucci, A., Kesterson,
R.A., Al-Obeidi, F.A., Hadley, M.E. and Cone, R.D.
(1995) Cyclic lactam -melanotropin analogues of
Ac-Nle4-cyclo[Asp5,D-Phe7, Lys10]
-melanocyte-stimulating hormone-(4-10)-NH2 with bulky
aromatic amino acids at position 7 show high antagonist potency and
selectivity at specific melanocortin receptors. J. Med.
Chem., 38,3454
-3461.[Web of Science][Medline]
Ingersoll, D.W., Morley, K.T., Benvenga, M. and Hands, C. (1986) An accessory sex gland aggression-promoting chemosignal in male mice. Behav. Neurosci.,100 , 777-782.[Web of Science][Medline]
Jones, R.B. and Nowell, N.W. (1973)
Effects of preputial and coagulating gland secretions upon aggressive
behaviour in male mice: a confirmation. J. Endocrinol.,59
, 203-204.
Kingsley, H.N. (1998) Disruption of the fifth melanocortin receptor alters inter-male aggression pheromones in house mice. Unpublished masters thesis, Department of Biology, University of North Carolina at Greensboro.
Labbe, O., Desarnaud, F., Eggerickx, D., Vassart, G. and Parmentier, M. (1994) Molecular cloning of a mouse melanocortin 5 receptor gene widely expressed in peripheral tissues.Biochemistry , 33,4543 -4549.[Medline]
McKinney, T.D. and Christian, J.J. (1970) Effect of preputialectomy on fighting behavior in mice. Proc. Soc. Exp. Biol. Med.,134 , 291-293.[Medline]
Morgan, C., Thomas, R.E., Ma, W., Cepoi,D., Novotny, M. and Cone, R.D. (1999) Decreased dominant behavior and pheromonal signaling in melanocortin type-5 receptor-deficient mice.Soc. Neurosci. Abstr. , 25, 243.1, p.605 .
Novotny, M., Harvey, S. and Jemiolo, B. (1990) Chemistry of male dominance in the house mouse, Mus domesticus. Experientia, 46,109 -113.[Web of Science][Medline]
Sandnabba, N.K. (1985) Differences in the capacity of male odors to affect investigatory behaviour and different urinary marking patterns in two strains of mice, selectively bred for high and low aggressiveness. Behav. Proc., 11,257 -267.
Sandnabba, N.K. (1986) Differences between two strains of mice, selectively bred for high and low aggressiveness, in the capacity of male odors to affect aggressive behavior. Aggress. Behav., 12,103 -110.[Web of Science]
SAS Institute (1995) JMP IN® 3.1.5 Windows Version. SAS Institute Inc., Cary, NC.
Tatro, J.B. and Reichlin, S. (1987)
Specific receptors for -melanocyte-stimulating hormone are widely
distributed in tissues of rodents. Endocrinology,121
, 1900-1907.
Thody, A.J. and Shuster, S. (1973) Possible role of MSH in the mammal. Nature,245 , 207-208.[Medline]
Accepted September 6, 2001
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  K. L.J Ellacott and R. D Cone The role of the central melanocortin system in the regulation of food intake and energy homeostasis: lessons from mouse models Phil Trans R Soc B, July 29, 2006; 361(1471): 1265 - 1274. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Morgan, R. E. Thomas, W. Ma, M. V. Novotny, and R. D. Cone Melanocortin-5 Receptor Deficiency Reduces a Pheromonal Signal for Aggression in Male Mice Chem Senses, February 1, 2004; 29(2): 111 - 115. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Kiyokawa, T. Kikusui, Y. Takeuchi, and Y. Mori Alarm Pheromones with Different Functions are Released from Different Regions of the Body Surface of Male Rats Chem Senses, January 1, 2004; 29(1): 35 - 40. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||