Altered sexual and social behaviors in trp2 mutant mice

Abstract

We have used gene targeting to generate mice with a homozygous deficiency in trp2, a cation channel expressed in the vomeronasal organ (VNO). Trp2 mutant animals reveal a striking reduction in the electrophysiological response to pheromones in the VNO, suggesting that trp2 plays a central role in mediating the pheromone response. These mutants therefore afford the opportunity to examine the role of the VNO in the generation of innate sexual and social behaviors in mice. Trp2 mutant males and nursing females are docile and fail to initiate aggressive attacks on intruder males. Male-female sexual behavior appears normal, but trp2 mutant males also vigorously mount other males. These results suggest that the cation channel trp2 is required in the VNO to detect male-specific pheromones that elicit aggressive behaviors and dictate the choice of sexual partners.

Figure 1

Strongly diminished responses in trp2−/− vomeronasal sensory neurons to synthetic and natural stimuli. (A) Representative EVG responses from wild-type (Upper) and trp2−/− mice (Lower). Responses were induced by 500-ms pulses of 2-heptanone at 10⁻⁷ M or 10⁻⁴ M, respectively. (B) Representative EVG responses from wild-type (Upper) and trp2−/− mice (Lower) induced by 500-ms pulses of urine diluted 1/10⁵ or 1/10², respectively. (C) Histograms showing collected results (mean ± SD) from +/+ (red), +/− (green), and −/− (blue) mice. Peak EVG responses from male and female animals were pooled. For stimulation with 10⁻⁷ M 2-heptanone, no responses are observed in trp2 mutants (n = 11). With 10⁻⁴ M 2-heptanone, responses are reduced to about 25% of control mice [+/+: 305 ± 66 μV (n = 7); trp2−/−: 77 ± 15 μV (n = 4); P < 0.0001]. For stimulation with urine diluted 1/10⁵, no responses are observed in trp2 mutants (n = 4). With urine diluted 1/10², responses are reduced to about 37% of control mice [+/+: 277 ± 61 μV (n = 6); trp2−/−: 103 ± 45 μV (n = 10); P < 0.0001]. Responses from +/+ and +/− mice were essentially identical (P = 0.21–0.80).

Figure 2

Trp2−/− males mate normally with females but display increased mounting toward other males. (A) Equivalent sexual behavior toward estrous females is exhibited by sexually naive trp2 (n = 11, blue bars) and control (n = 12, red bars) male mice in a 30-min mating test. (B) Increased intermale mounting is observed in trp2−/− mice (n = 23) in comparison to controls (n = 25) upon the addition of a male to their home cage in a 15-min test. Both before and after mating experience with females, trp2 males exhibited a significantly greater number of intermale mounts than controls (mount frequency, P < 0.001; Mann–Whitney U test). In fact, trp2−/− mice displayed a significant increase in intermale mounting after sexual experience with females (mount frequency, P < 0.05, Wilcoxon signed rank test). (C) trp2−/− males continue to mount males at a high rate even when a female is present. Sexually experienced trp2−/− (n = 12) and control (n = 12) male mice were observed for behaviors exhibited upon the simultaneous addition of a male and nonestrous female mouse into their home cage in a 15-min test. Trp2 males exhibit a 2-fold preference for mounting females, whereas in the control group there is a 10-fold preference for mounting females.

Figure 3

Lack of aggressive behavior in trp2−/− males and lactating females. (A) trp2−/− males are not aggressive in a resident–intruder assay. Trp2 (n = 23, blue bars) and control (n = 25, red bars) male mice were observed for behaviors elicited by the addition of a male to their home cage for 15 min prior to and after mating experience with females. Trp2 males were significantly different from control males by all measures of aggressive behavior (latency to attack, attack frequency and attack duration, P < 0.0001, Mann–Whitney U test). There was also a significant increase in all parameters of aggressive behavior after sexual experience in the control group (P < 0.05, Wilcoxon signed rank test), but sexual experience did not stimulate aggression in the trp2−/− group. (B) trp2−/− lactating females are not aggressive in a maternal aggression assay. Lactating trp2 (n = 11) and control (n = 11) mothers were tested for aggression toward intruder males in a 15-min test. Trp2 females were significantly different from control females by all measures of aggressive behavior (P < 0.05). Forty-five percent of control females responded aggressively toward intruder males on at least one of the test sessions, whereas trp2 females never initiated attacks.

Figure 4

trp2−/− males become subordinate to control males and exhibit altered territorial behavior when paired with other trp2 males. (A) trp2−/− and control males mark in a dominant fashion before fighting experience with each other. trp2−/− (n = 11) and control (n = 11) males, which had been singly housed, were placed on opposite sides of a wire mesh barrier in a two-chamber cage lined with filter paper for 30 min. The total number of marks by controls (89.8 ± 20.7) and mutants (91.1 ± 24.0) did not significantly differ. (B) trp2−/− males become subordinated and suppress marking after repetitive pairing with controls (n = 11 pairs). In 10 of 11 of cases the control was dominant over the trp2 mutant, as assessed by marking. (C) trp2−/− males do not form normal dominant–subordinate relationships. After repetitive pairing as in B, analysis of the marking patterns of mutant pairs (n = 6 pairs) reveals that in all cases, neither member of the pair suppressed marking behavior in the presence of the other. In contrast, analysis of marking patterns of control pairs (n = 5 pairs, not shown) demonstrated that in all cases one of the males became dominant over the other.