Effects of an Auditory Species Typical Cue on the Conditioning of Male & Female Japanese Quail (Coturnix japonica) By Carlina Muglia

Transcription

1 Effects of an Auditory Species Typical Cue on the Conditioning of Male & Female Japanese Quail (Coturnix japonica) By Carlina Muglia Abstract This study examined if there was a difference between conditioned sexual responding produced by an arbitrary and a species typical conditioned stimulus (CS) in male and female Japanese quail (Coturnix japonica). Male and female quail were exposed to conditioning trials in which playback of either a male s crow or an arbitrary tone signaled the beginning of a five minute copulatory opportunity. There was a marginally significant difference in the frequency with which birds were squatting between the paired and unpaired conditions and a significant difference in the duration with which the birds were squatting between the paired and unpaired conditions. There was also a significant difference in the number of fertile eggs laid in the paired and unpaired conditions during conditioning. All other variables for the CS period yielded non significant results. Data from the unconditioned stimulus (US) periods has yet to be scored and analyzed. Therefore, the effectiveness of the call in conditioning behavioral changes in males and females is as yet ambiguous. In the classical model of Pavlovian conditioning, an arbitrary stimulus (the conditioned stimulus or CS) is presented immediately following a biologically relevant stimulus (the unconditioned stimulus or US) that elicits a reflexive behavior (the unconditioned response or UR). If the CS US pairing is repeated, the CS will begin to elicit responses that are related to the reflexive behavior previously shown in response to the US. For example, Pavlov (1927) conditioned dogs to salivate to the clicking sound of a metronome (the CS) by pairing it with the presentation of food (the US). Sexual conditioning is a specialized form of Pavlovian conditioning in which an arbitrary CS is repeatedly paired with a copulatory opportunity that serves as the US. If an association between the CS and US is learned, the animal will come to direct sexual responses toward the CS (Cusato & Domjan, 1998, 2000; Hollis, Cadieux, & Colbert, 1989; Krause, Cusato, & Domjan, 2003). Copulatory interactions have a wide degree of variation between sexes, species, and mating systems, and differ seasonally and 1

2 developmentally, which also makes them dissimilar to more traditional USs (Crawford, Holloway, & Domjan, 1993). A popular species for laboratory investigations of sexual conditioning is the Japanese quail (Coturnix japonica). Early studies focused in particular on the more overt sexual behaviors of the male of this species, but more recently have begun to examine the effects of sexual conditioning on female behavior. For example, Gutiérrez & Domjan (1997) compared the effects of sexual conditioning on the behavior of male and female Japanese quail. The investigation illustrated a marked difference between the behavioral manifestations of sexual learning in male and female quail; the former showing strong approach behavior directed toward the CS and the latter showing equally vigorous squatting behavior in the US. Female quail demonstrate sexual receptivity by squatting, and through this behavior, control the male copulatory behaviors of latency, frequency, and efficiency of grabs, mounts, and cloacal contacts. Domjan, Mahometa, & Mills (2003) suggested that by controlling the male s behavior in the aforementioned manner, the female is indirectly able to control the rate at which her eggs are fertilized and is therefore able to exert control over sexual encounters. These findings are contrary to past research which suggests a considerably smaller role for females since their sexually conditioned behavior is often more subtle, unlike the aggressive approach behaviors of the males (Domajn & Hall, 1986b; Gutiérrez & Domjan, 1997). Much of the existing literature on sexual conditioning focuses on the use of arbitrary lights or tones as the CS (e.g., Domjan, Lyons, North, & Bruell, 1986; Gutiérrez & Domjan, 1997; Hollis et. al., 1989; Mahometa & Domjan, 2005). Mahometa & Domjan (2005) were able to sexually condition an increase in fertility in Japanese quail by signaling both members of a mating pair with an arbitrary visual stimulus. A study by Davis (2004) yielded similar results 2

3 using an arbitrary 1000Hz tone. Additionally, conditioning both members of the mating pair overcame fertility deficits. Subthreshold fertility increases in both the male and female may explain why both members of the mating pair must be signaled in order to obtain an overall increase in fertility. These subthreshold increases may combine to form a summation effect resulting in an overall increase. A possible way to overcome the inefficiency of having to signal both members of the mating pair is to use a CS that is more salient. In the early 1990s, researchers such as Domjan (1994, 1998) and Timberlake (1993) began to examine the possibility of using ecologically relevant species typical stimuli in the context of behavior systems. The overall conclusions were that such stimuli would alleviate one of the harshest criticisms of learning research that laboratory conditions and stimuli were not easily generalized to naturalistic settings. It was also concluded that a greater scope of knowledge could possibly be gained by taking such an ecological approach. Since then, experiments have successfully conditioned scent and flavor preferences in hamsters using other hamsters as CSs in a visual and social manifestation (Vasileva, Syau Cheng, Ekaterina, & Johnston, 2001; Lupfer, Frieman, & Coonfield, 2003). Most relevant to the current study, however, Cusato & Domjan (1998) explored the functional differences between species typical and arbitrary cues as CSs by using an ecologically relevant visual cue and an arbitrary cue in a sexual conditioning procedure with Japanese quail. Speciestypical visual cues effectively increased the males approach and copulatory responses in comparison to the arbitrary CSs. The functional significance of the unmated male Japanese quail s call has been subject to speculation. It is possible that the call serves the purpose of spacing males territorially, for defense, or for attracting females (Potash, 1974). Goodson & Adkins Regan (1997) found that 3

4 female Japanese quail maintained an overall shorter distance from a speaker playing the male s crow than from a speaker playing a reversed crow, and that males were indifferent to either call. These findings are consistent with the possible explanation of the purpose of the male s call in Potash (1974). The current study was conducted to more fully assess the efficacy of the male Japanese quail s call in sexual conditioning by examining its effects on the behavior and fertility of both male and female quail. The variables manipulated in the current study were: Sex (male and female), the type of auditory cue (1000Hz tone and male crow) and pairing (paired and unpaired). Fertility data was gathered by collecting eggs laid by each of the females daily during conditioning, and behavioral data was collected by taking an analog video recording of every third day of conditioning for each subject in a 2 (male vs. female) X 2 (species typical vs. arbitrary) X 2 (paired vs. unpaired) between subjects design. This study was conducted in order to determine if the male s call was effective as a CS. It was hypothesized that females in the paired species typical group would have significantly different sexually conditioned behavior than those in the paired arbitrary group, unpaired species typical, and unpaired arbitrary groups, and that the difference would manifest itself in receptivity or squatting behavior and percentage of fertile eggs laid. It was also hypothesized that the males in the paired species typical group would have a significantly different change in sexually conditioned behavior compared to the males in the paired arbitrary, unpaired species typical, and arbitrary groups, and that these differences would manifest themselves in goal tracking behavior and fertility. 4

5 Method Subject Subjects were twenty eight sexually mature male and twenty seven sexually mature female Japanese quail (Coturnix japonica). All birds were housed in individual cages in a climate controlled environment operating on a 16:8 hour on:off extended light cycle to ensure copulation. The females were housed in a separate sound isolated room in order to eliminate extraneous exposure to the call in non conditioning contexts. Female egg laying was monitored prior to the start of the experiment. Only females that had consistent egg laying (approximately 1 egg per day) were chosen as subjects for this study. All males were given a five minute copulation test in order to examine their ability to complete a grab, mount, and cloacal contact cycle. Only males that were able to complete the cycle properly were chosen for this study. All birds were allowed continuous access to food and water. Apparatus Seven identical test cages were used as the experimental chambers. Each cage was 120cm (wide) x 65cm (high) x 105cm (deep) and made of sealed plywood. Each chamber had a wall separating it into two unequal compartments. The left compartment of each chamber served as the context in which the CS was presented and in which the US took place. Its dimensions were 65cm (wide) x 65cm (high) x 105cm (deep). The right compartment served as a holding chamber for the stimulus males and females. This side measured 45cm (wide) x 65cm (high) x 105cm (deep). Both sides of each chamber had a chicken wire mesh and sealed plywood door to provide access to the chamber and ease of recording. In the center of each dividing wall between the subject and stimulus compartments of each chamber was a 15cm (wide) x 15cm (high) vertical sliding door that, when open, allowed one way access to the subject compartment from 5

6 that of the stimulus bird. Each sliding door was made of plexi glass and had black mesh attached to the stimulus side. The lights on the stimulus side of the compartment were shut off completely and the front door was covered in black plastic in order to prevent light from entering the chamber. This was done in order to create a two way mirror effect. When a female was in the subject chamber she was unable to see into the darkened male compartment. The male, however, was able to see the female in the lighted side of the chamber. In order to ensure the male maintained visual access to the female, each male s movement was restricted to a 270cm 2 area by a 11cm (high) x 18cm (wide) x 15cm (deep) triangle made of wood and covered with chicken wire. This was done to prevent the males from crowing, which is done by keeping them in visual contact with a female. This arrangement was emulated with the female stimulus birds as well, in order to have consistency within the experiment. A stimulus speaker was located 51cm from the front and back of the cage, 38cm from the top of the cage and 5cm from the floor of the cage on the left wall of the female compartment. The flooring consisted of coated wire mesh and was divided into fifteen 20cm x 20cm zones that were used to assess female ambulatory behavior and male sign and goal tracking behavior. Prior to the start of the experiment, several male calls were recorded using an Olympus DS 2 Digital Voice Recorder. The files were transferred into Audacity (v.1.2.1) on an Apple Mac OSX computer for audio editing. Each call was compared to sonograms from previous research that represented the typical unmated Japanese quail crow (Goodson & Adkins Regan, 1997; Potash, 1974; Shaw & Kennedy, 2002). The two recordings that most closely resembled the sonograms were digitally mastered and recorded to a compact disc at the rate of one crow every five seconds for thirty seconds, alternating between the two crows. An arbitrary 1000Hz tone was also recorded to the same compact disc as a separate track using the same intervals; one 6

7 second of tone every five seconds for a total of thirty seconds. The 1000Hz tone was chosen because it falls within the quail auditory range based on inferences drawn from the quail vocal range, kHz (Mills, Crawford, Domjan, & Faure, 1997). Recording for behavioral data utilized three Security Labs CCD video cameras that transferred data to a Canopus ADVC 100 analog recorder, as well as to videocassette. Postconditioning, the analog recordings were transferred to an Apple Mac OSX computer and converted to digital recordings using Discreet Cleaner 6.01 for easier scoring. All scoring took place on a Dell Optiplex Gx50 computer on a Microsoft Office 2000 system and utilized the Noldus Information Technologies Observer 4.1 program. Data was analyzed in SAS Institute Inc. s Statview for Windows v Procedure Prior to the beginning of the experiment, the male and female quail were assigned to one of four squads (Squad A, Squad B, Squad C, or Squad D). In Squads A and B males were the subjects and females were stimulus birds, and in Squads C and D females were the subjects and males were the stimulus birds. Additionally, these squads were assigned a call type. Squads A and C received the species typical cue and Squads B and D received the arbitrary cue. Each group was finally divided into two further groups per squad (ns = 3 or 4), either paired or unpaired. This created eight groups. Four each in the male and female condition: species typical paired (STP), species typical unpaired (STU), arbitrary paired (AP), and arbitrary unpaired (AU). The arbitrary groups received the 1000Hz tone recording and the species typical groups received the recording of the male s crow. In the paired conditions the subjects were exposed to the thirty second CS, immediately after which, the door between the subject and stimulus compartments was opened, allowing the stimulus access to enter the subject s compartment. If 7

8 the stimulus did not enter on its own within thirty seconds, it was pushed through the doorway. After the stimulus entered the subject s chamber, US time began. The birds were allowed a five minute copulation period. After five minutes the stimulus was removed. In the unpaired groups the subjects received the same thirty second CS followed by a forty five minute latency prior to the start of the five minute US. After the forty five minute latency, the center door was opened and the stimulus entered or was pushed into the experimental chamber within thirty seconds, and a five minute US period began. After the US period, the stimulus was removed from the cage. Twenty one days of conditioning were conducted, as well as a ten day clean out period. The physiology of the female quail reproductive system is such that it can hold sperm from a single sexual encounter for up to ten days. The clean out period allowed excess sperm from conditioning to be expelled. Clean out is typically ten to twenty one days long and is conducted in the period immediately following conditioning and immediately preceding the post test. This ensures that any fertile eggs laid by the females are the result of sexual interaction during the post test and not during conditioning. The conditioning trials were scored for the number of times the female subjects crossed into a new zone (line crosses), for the amount of time the male subjects spent within 38cm of the speaker or center door (sign and goal tracking), and for the frequency and length of time the females spent squatting (squat frequency and squat duration). A female was considered to be squatting if she bent her legs and lowered her body, so that it came into contact with the floor. Fertility was measured in terms of the total number of eggs laid, total number of fertile eggs, and the percentage of fertile eggs laid. Difference scores are those in which the values of the first day of data collection are subtracted from each day in order to control for initial variability between subjects in all of the behavioral measures. 8

9 Results Difference scores for line crosses, squat frequency and duration, sign tracking, and goal tracking from the CS period were evaluated using five separate analyses of variance (ANOVAs) in which the day of conditioning served as a within subjects variable, and the CS type and pairing, and sex, served as between subjects variables. For the line crossing measure, the main effect of Pairing was not significant [F(1, 27) = 2.591, p =.1205], and the interactions between the day of conditioning and Pairing, Call Type, and the three way interaction between Pairing, Call Type, and day of conditioning were not significant [Fs(3, 27) = 1.327, 2.280, and 2.427, respectively, ps >.05]. The difference score analysis for this data yielded the same results for the interactions between the day of conditioning and Pairing, day of conditioning and Call Type, and the three way interaction between Pairing, Call Type, and day of conditioning [Fs(3, 27) = 1.327, 2.280, and 2.427, respectively, ps >.05]. The main effect for Calling in the squat frequency measure was significant [F(1, 27) = 3.429, p <.05]. When difference scores for this data were run, the significant difference disappeared. The squat frequency measure had main effects of Pairing and Call Type that were non significant [Fs(1, 27) = and 1.244, respectively, ps >.05] as was the interaction between day of conditioning and Call Type [F(3, 27) = 1.611, p >.05]. Squat duration yielded two significant main effects of Pairing and Call Type [Fs(1, 27) = and 9.555, respectively, ps <.05] (see Figure 1 for Pairing data). There was a non significant interaction between the day of conditioning and Pairing [F(3, 27) = 1.221, p >.05]. When difference scores were run on this data, the main effect for Pairing was not significant along with the interaction between the day of conditioning and Pairing [Fs(1, 27) = and 1.221, respectively, ps >.05]. The analysis for Goal Tracking yielded a non significant interaction between Call Type and Pairing [F(1,27) = 1.247, p >.05]. The difference 9

10 score analysis yielded a non significant main effect of Pairing and interaction between Pairing and Call Type [Fs(1, 27) = and 1.197, respectively, ps >.05. The Sign Tracking measure yielded a non significant main effect of Call Type [F(1, 27) = 1.525, p >.05] and interactions between Pairing and Call Type, and day of conditioning and Pairing [Fs(3, 27) = and 1.573, respectively, ps >.05]. In the difference score analysis the main effect of Pairing and the interaction between day of conditioning and Pairing were non significant [Fs(3, 27) = and 1.573, respectively, ps >.05]. No other behavioral measures had significant main effects or interactions (Fs < 1). Additional ANOVAs were run on fertility data from conditioning. There was one significant main effect of Pairing in conditioning percentage of fertility [F(1, 27) = , p =.0039] (see Figure 2) and a marginally significant main effect of Pairing in conditioning fertility [F(1, 27) = 3.139, p =.0925]. No other fertility measures had significant main effects or interactions (Fs < 1). 10

11 Discussion This study was conducted in order to determine if there was a difference in the conditioned effects of the male s call in comparison to an arbitrary CS and to see if there was a difference between the sexually conditioned behavior of the male and female quail to these stimuli using a 2 (male vs. female) X 2 (species typical vs. arbitrary) X 2 (paired vs. unpaired) between subjects design. The significant data for the squatting behavior during the CS is consistent with the hypotheses for this particular experiment. It is however contrary to previous findings, such as Gutiérrez & Domjan (1997), which showed no behavioral changes in the females during the CS period of conditioning, but showed an increase in receptivity during the US period in the paired groups in comparison to the unpaired groups. It is possible that the results for the US will be replicated with further scoring and analysis. The significant main effect of percentage of fertility demonstrated in conditioning also leads one to believe that a significant change in squatting would have continued into the US period. Goodson & Adkins Regan (1997) indicated that females show goal tracking behavior. In that particular study, the criterion for approach to the speaker broadcasting the male s crow was 1m. In this study, during the male CS period, the criterion used was 38cm. This may account for the lack of goal sign and goal tracking behavior seen in the males. It is possible that the criterion in this experiment was too stringent. In order to be able to interpret the results of this study to the fullest extent, data from the US period and Post Test must first be analyzed. Current findings suggest that there could be significant behavioral differences in the US that are driving the significant main effect of fertility in conditioning. Taking the current findings alone, one is able to conclude that a species typical cue is at least as effective as an arbitrary cue at conditioning increases in fertility because the 11

12 birds were able to use the cue as a predictor of sex. In the future it is necessary to complete the analyses of the US period. It is also important to look at a variety of noise cues created by the male to see if perhaps there are more salient calls than that of the unmated male. It may also be prudent to examine a different presentation of the male s crow. In its current form it follows a very unnatural pattern, which may do more to confuse the subjects than to reflect a naturalistic experience. 12

13 References Adkins, E.K., Boop, J.J., Koutnik, D.L., Morris, J.B., & Pniewski, E.E. (1980). Further evidence that androgen aromatization is essential for the activation of copulation in male quail. Physiology and Behavior, 24, Adkins, E.K. (1981). Hormone specificity, androgen metabolism, and social behavior. American Zoologist, 21, Cusato, B., & Domjan, M. (1998). Special efficacy of sexual conditioned stimuli that include species typical cues: Tests with a CS pre exposure design. Learning and Motivation, 29, Cusato, B., & Domjan, M. (2000). Facilitation of appetitive conditioning with naturalistic conditioned stimuli: CS and US Factors. Animal Learning and Behavior, 28, Crawford, L., Holloway, K., & Domjan, M. (1993). The nature of sexual reinforcement. Journal of the Experimental Analysis of Behavior, 60, Davis, L., & Cusato, B. (2004). Can Pavlovian conditioning overcome the negative effects of stress? Fertility effects in female Japanese quail (Coturnix japonica). Paper presented at the Conference of the Southwestern Comparative Psychological Association. San Antonio, Tx. Domjan, M., & Hall, S. (1986b). Determinants of social proximity in Japanese quail (Coturnix japonica): Male behavior. Journal of Comparative Psychology, 100, Domjan, M., Lyons, R., North, N.C., Bruell, J. (1986). Sexual Pavlovian conditioned approach behavior in male Japanese quail (Coturnix coturnix japonica). Journal of Comparative Psychology, 100, Domjan, M. (1994). Formulation of a behavior system for sexual conditioning. Psychonomic Bulletin & Review, 1, Domjan, M. (1998). Going wild in the laboratory: Learning about species typical cues. In D. L. Medin (Ed.), The Psychology of Learning and Motivation (vol. 38, pp ). San Diego: Academic Press. Domjan, M., Mahometa, M.J., & Mills, A.D. (2003). Relative contributions of the male and female to sexual behavior and reproductive success in the domesticated quail (Coturnix japonica). Journal of Comparative Psychology, 117, Goodson, J.L., & Adkins Regan, E. (1997). Playback of crows of male Japanese quail elicits female phonotaxis. Condor, 99,

14 Gutiérrez, G., & Domjan, M. (1997). Differences in the sexual conditioned behavior of male and female Japanese quail (Coturnix japonica). Journal of Comparative Psychology, 111, Hollis, K.L., Cadieux, E.L., & Colbert, M.M. (1989). The biological function of Pavlovian conditioning: A mechanism for mating success in blue gourami (Trichogaster trichopterus). Journal of Comparative Psychology, 103, Krause, M.A., Cusato, B., & Domjan, M. (2003). Extinction of conditioned sexual responses in male Japanese quail (Coturnix japonica): Role of species typical cues. Journal of Comparative Psychology, 117, Lupfer, G., Frieman, J., & Coonfield, D (2003). Social transmission of flavor preferences in two species of hamsters (Mesocricetus auratus) and (Phodopus campbelli). Journal of Comparative Psychology, 117, Mahometa, M.J., & Domjan, M. (2005). Classical conditioning increases reproductive success in Japanese quail (Coturnix japonica). Animal Behaviour, 69, Mills, A.D., Crawford, L.L., Domjan, M., & Faure, J.M. (1997). The behavior of the Japanese or domestic quail, Coturnix japonica. Neuroscience and Biobehavioral Reviews, 21, Pavlov, I.P. (1927). Conditioned reflexes. (G.V. Anrep, Ed. & Trans.). London: University Press. Oxford Potash, L.M. (1974). An experimental analysis of the use of location calls by Japanese quail, Coturnix japonica. Behaviour, 54, Shaw, B.K., & Kennedy, G.G. (2002). Evidence for species differences in the pattern of androgen receptor distribution in relation to species differences in an androgen dependent behavior. Journal of Neurobiology, 52, Vasileva, N.Y., Lai, S., Petrova, E.K., Johnston, R. E. (2001). Development of species preferences in two hamsters, Phodopus campbelli and Phodopus sungorus: Effects of cross fostering. Ethology, 107,

15 Figure Captions Figure 1. Duration of squatting during the CS as a function of time measured in seconds (STU, STP, AU, AP). Figure2. Percentage of fertile eggs laid during conditioning between the subject and stimulus birds 15

16 Squat Duration between Paired and Unpaired Groups paired unpaired Pairing 16

17 % Fertility During Conditioning Paired Stimulus UnPaired Stimulus Paired Subject Unpaired Subject Paired Stimulus UnPaired Stimulus Paired Subject UnPaired Subject Bird Type & Pairing 17