1. Question--Last Lecture (Chp. 6) Two main alternative reproductive strategies in male bluegill fish: (heritable) territorial male: – large colorful males that defends territories and solicits females to release eggs so that he can fertilize the eggs –territorial males will give rise to young that are large and brightly colored sneaker males that becomes a female mimic: –sneaker males “sneak” matings when the large territorial male is preoccupied –sneaker males are younger and smaller and possess a light body coloration –as sneaker males age they become female mimics, their body size increases and they develop a coloration pattern with dark vertical bars (like females) –female mimics (large drab males) look like females and they hang around in the vicinity of real females –female mimics are ready to release sperm when the female releases her eggs in response to the behavior of the territorial male

2. Chp 7: Hormonal Influences on Courtship Behavior Overview: Courtship tends to be a sexually dimorphic response: –in many species, courtship is initiated largely by males –links to female preferences and male’s comparative quality Ex. Production of EODs in Weakly Electric Fish: –fish communicate via electric organ discharges (EODs) –gonadal steroids influence the display of the EOD and of “chirping”--the male’s courtship response Ex. Production of the “mating call’ in Xenopus (clawed frog) –the ‘mating call’ is used to attract females to a breeding area –gonadal steroids act to organize and activate the ‘mating call’ Ex. Production of “song” by song birds –males produce song to attract females for mating –gonadal steroids act to organize and activate both learning and production of song

3. Courtship As a Sexually Dimorphic Response Courtship tends to be a sexually dimorphic response. For example: In most fish species, males do all or the majority of “chirping.”In most fish species, males do all or the majority of “chirping.” In frogs like Xenopus, only males can produce the “mating call.”In frogs like Xenopus, only males can produce the “mating call.” In several bird species, only males produce song. Why are courtship responses sexually dimorphic? Trivers (1985) suggests that sex differences in courtship behavior reflect differences in the relative investment of energy and time in reproduction. –females spend more time and energy –males spend less time and energy

4. Investment of energy & time in reproduction: Energetic investment in production of gametes Produce sperm, which consist of male’s chromosomes surrounded by a protein coat and flagellum Produce eggs, which consist of female’s chromosomes and yolk, a high energy nutritive substance Males Females “cheap” “expensive” Number of gametes produced over a lifetime Produce large numbers of sperm Produce small numbers of eggs Ex. humans (men), 1 ejaculation can release 100 million sperm Ex. humans (women), over lifetime, see production of a few hundred eggs

5. Investment of energy & time in reproduction: Care of offspring Less time and energy is devoted in care of young Much greater time and energy is devoted to care of young Males Females development in utero (gestation) feeding and caring for young once born or hatched

6. Why is courtship largely initiated by males? female is limited by costs of making and rearing young female maximizes her reproductive investment by choosing to which males will fertilize her eggs males must demonstrate their comparative quality To mate with females: winning territorial battles showing specific courtship responses possessing other traits (e.g., colorful plumage) “Good” male: male with high genetic quality (good genes) male that cares for young (by feeding or protection)

7. Courtship in the Weakly Electric Fish Weakly electric fish communicate and navigate by use of electrical signals. electrical signals are called electric organ discharges (EODs) EOD is produced by the electric organ located in the tail –EOD is spontaneously produced (important for navigation in murky waters and for identification of others in environment) –EOD is also produced in response to cues that trigger mating: rain triggers the production of “chirping” by males (courtship response) EOD is perceived by specific sensory cells called electroreceptors –electroreceptors are concentrated in the head region –transduce electrical stimuli into chemical stimuli to activate sensory pathways in brain

8. Courtship in the Weakly Electric Fish The electric organ is involved in producing EOD: neurons within the hindbrain control production of spontaneous EODs –pacemaker nucleus-->electromotor neurons in spinal cord-->electric organ –electric organ consists of modified muscles cells called electrocytes that produce the EOD neurons in forebrain control production of EODs in response to various cues (e.g., EODs produced by females or to activating cues like “rain”) –prepacemaker nucleus-->pacemaker nucleus will stimulate production of EODs The EOD varies in “form”: discharge frequency: number of pulses produced per given length of time; frequency discharge can be low or high pulse duration: the waveform may be long or short there are species differences and sex differences in the form of the EOD

9. Courtship in the Weakly Electric Fish Spontaneous EODs provide information about sex and age of individual: mature males: –low discharge frequency (50-90 Hz) –long pulse duration mature females: –high discharge frequency (100-150 Hz) –short pulse duration juveniles: –intermediate values for both mature males mature females

10. Courtship in the Weakly Electric Fish Gonadal steroids influence discharge frequency and pulse duration of EOD: androgens act to broaden pulse duration and lower discharge frequency estrogen acts to increase frequency discharge Male Female EOD of low frequency and long duration EOD of high frequency and short duration castrate male increase in frequency and decrease in duration ovariectomize female decrease in frequency further decrease in frequency and increase in pulse duration [Juvenile] administer androgens

11. Courtship in the Weakly Electric Fish Gonadal steroids mediate their effects in two ways: androgens alter pulse duration by acting at electric organ –electrocytes possess androgen receptors –androgens act at electrocytes to increase thickness of the electrocyte membrane –a thicker membrane can store more electrical charges that can then be released for a longer period of time –androgens “prolong EOD waveform” (long pulse duration) by increasing the thickness of the electrocyte membrane androgens and estrogens are believed to alter discharge frequency by acting on groups of neurons within the brain –androgens act to decrease discharge frequency –estrogen acts to increase discharge frequency

12. Courtship in the Weakly Electric Fish Courtship response of males is called “chirping”: brief and rapid increase in EOD frequency important in stimulating a female to lay eggs Ex. Observations made by Hagedorn: –“The night of spawning is an electrical extravaganza, males will fight for many nights for [dominance in the mating hierarchy], the females defend spawning territories (floating plants) and the dominant male will spawn only with the dominant female. The female hangs almost vertically in the plants, while the male courts her with a barrage of [electrical] “chirping” (60 to 80 chirps/minute). As the female lays her eggs, she produces low amplitude chirps, then the male rubs through the plants fertilizing the eggs. Peripheral, subordinate females try to sneak their eggs into the spawning territory [often by turning off their EOD as they rush towards the plants], only to be driven away by the dominant pair.”

13. Vocal Courtship in Frogs Male frogs typically produce the majority of vocalizations. Vocal communication provides a selective advantage in these species: most frogs live in a muddy and wet environment, and they are behaviorally active at night (nocturnal) as a consequence, visual cues would not be very helpful in finding a mate sounds have the advantage of traveling long distances and around obstacles, and they do not need to be in a certain orientation for effective communication to occur The vocalizations serve two main functions: attract females for mating--production of “mate calling” defend territorial boundaries (aggressive response)

14. Vocal Courtship in Frogs The “call” of the frog contains important information: sound amplitude (loudness) and sound frequency (pitch) –sound amplitude and frequency changes as the frog ages –frogs exhibit indeterminate growth--”as they get older they get bigger” –an old frog will have a larger vocal apparatus-->he will produce a louder sound at a lower frequency duration of call (long or short): indication of health overall-pattern of the call: indicates species of male calling (viability of young) You are “it” as a male if you are large, and old and call for a long duration! females prefer to mate with large, old males that call for a long period of time: –these males have successfully survived their environment and are healthy –consequently, these male must possess “good genes” that will increase the likelihood that their young will survive

15. Vocal Courtship in Frogs There are two types of calls that can be made: “mate calling” –this is exclusively produced by males (courtship response) –typically, clawed frogs are dormant during the dry season, but become active when the rainy season starts--“rain” activate courtship response –serves to attract a female to breeding area and to arouse the female (stimulates her to “wiggle around in the mud”) –mate calling will lead to mating in which the male clasps the female from behind and then fertilizes eggs released by the female ticking –can be produced by males or females –serves to release a female when she is not receptive –in the mud, males can also clasp other males; consequently, males also “tick” to gain release

16. Vocal Courtship in Frogs Gonadal steroids have both organizational and activational effects on vocalizations: organizational effect: androgens have a masculinizing effect on laryngeal muscles that leads to masculinization of brain nuclei (similar to SNB--target derived neurotrophic function) –males have laryngeal muscles (that control the larynx) with many muscle fibers that can contract and relax at a fast rate (100 contractions/sec) –in contrast, females have laryngeal muscles with fewer muscle fibers that contract and relax more slowly (40 contractions/sec) –as a consequence, males have more motorneurons that innervate laryngeal muscles (motorneurons within cranial nerve nuclei--IX and X) –in addition, there are more forebrain connections to the dorsal tegmental area of the medulla (DTAM) that projects to cranial nerve nuclei to control vocalization –DTAM (and its connections to cranial nerve nuclei IX and X) is believed to be responsible for producing the “mating call”

17. Vocal Courtship in Frogs Comparison between males and females: forebrain DTAM cranial nerve nuclei IX & X Males laryngeal muscles “mate calling” forebrain DTAM Females laryngeal muscles no “mate calling” cranial nerve nuclei IX & X Males have more and faster laryngeal muscles, more neurons in cranial nerve nuclei IX and X, and more connections within the brain to mediate “mate calling.” Receptors for gonadal steroids are present at each level within brain and within laryngeal muscles.

18. Vocal Courtship in Frogs Gonadal steroids have both organizational and activational effects on vocalizations: activational effect: androgens have an activational effect on the production of “mate calling” normal adult males castrate will not show “mate calling” administer exogenous androgens will show “mate calling” What would happen if you gave adult females androgens?

19. Vocal Courtship in Frogs Summary: giving adult females androgens would not stimulate “mate calling”--why? –the females lack the organizational effects of androgens during development that masculinize the laryngeal muscles first and then the brain androgens act early in development to organize the laryngeal muscles and brain in a masculine fashion the organizing effects of androgens then set-the-stage for androgens to activate the brain (permissively) to show courtship behavior and mating when the opportunity arises

21. Vocal Courtship in Songbirds Song is used for communication among birds: courtship responses (attracting a mate for mating) aggressive responses (protecting territorial boundaries) Species differences in the production of song: in many species, the male produces all or the majority of song –Ex. zebra finch and canary –clear link between male’s display of song as a courtship behavior in other species, both males and females can sing –Ex. white-browed robin chat in which both males and females routinely sing in duets, although females have a smaller song repertoire –Ex. bay wren in which both males and females routinely sing in duets, with no sex difference in the size of song repertoire

22. Vocal Courtship in Songbirds Distinctions can be made among seasonal and nonseasonal breeders: nonseasonal breeders: –song production and mating occur throughout the year –food and nesting materials are available throughout the year –gonadal steroids are involved in organizing the adult male songbird brain in a masculine fashion and play an important but permissive role in the adult seasonal breeders: –song production and mating occur at specific times of the year –gonadal steroids are important for “activating” the display of song behavior and mating –gonadal steroids are also important for organizing the adult male songbird brain in a masculine fashion –seasonal changes in gonadal steroids and the display of song behavior and mating are reflected in seasonal changes in brain structure and connections

23. Vocal Courtship in Songbirds Song: definition: series of syllables, or brief sounds, arranged in a rhythmic sequence a given song can last from a few seconds to minutes different species of birds produce different songs (species-specificity) A songbird must “learn” how to sing. There are 3 stages to song learning and song production: –acquisition phase (sensorimotor learning) and a production phase (motor response) –subsong: considered “bird-babble”--male produce crude sounds that are highly variably in structure (canary--40 days of age)--acquisition –plastic song: bird starts to produce more polished sounds that begin to resemble a song (canary--60 days of age)--acquisition –crystallized song: well-defined song that is invariant in structure, can see clearly defined notes and motifs (canary--240 days of age--8 months)--production

24. Vocal Courtship in Songbirds Song learning and production require 2 main factors: ability to hear (hearing songs of others and own song) presence of gonadal steroids Hearing: if a bird is deafened (cannot hear) while he is acquiring song (during subsong or plastic song), he will not be able to produce normal song if a bird is deafened after he has acquired crystallized song, then he will be able to produce normal song--this ability depends on how he learns –age-limited learner: male goes through 3 stages of song learning and production and then song stabilizes; deafening after production of crystallized song has no major effect –open-ended learner: males goes through 3 stages of song learning during first breeding season, and then repeats each phase during subsequent breeding seasons (learning new song syllables); deafening after production of crystallized song has no effect during current breeding season but will drastically reduce ability during subsequent seasons

25. Vocal Courtship in Songbirds Gonadal steroids: gonadal steroids play a role in the both the organization and activation of song behavior Activational effects: gonadal steroids increase the production of song behavior (performance) both metabolites of testosterone (estrogen and DHT) are necessary for full response Ex. adult male zebra finch adult male zebra finch castrate reduction in rate of song production replace testosterone increase in song production

26. Vocal Courtship in Songbirds Activational effects: gonadal steroids also play a role in the acquisition of song behavior testosterone is important for acquisition of crystallized song estrogen (from nontesticular sources) is believed to be important for acquisition of subsong and plastic song Ex. swamp sparrow: males must hear song between 20-60 days after hatching, they will produce subsong (8 mo), plastic song (10 mo), & crystallized song (12 mo) young male swamp sparrows (3 weeks) castrate plus expose them to song [low T, high E] males produce subsong & plastic song, but not crystallized song administer exogenous testosterone crystallized song

27. Vocal Courtship in Songbirds Organizational Effects: gonadal steroids play a role in organizing the male brain to produce song as an adult in response to elevated levels of these hormones several brain areas involved in either sensorimotor learning or the production of song are masculinized in the male: –MAN - magnocellular nucleus of the anterior neostriatum –HVc - “high vocal center”, AKA hyperstriatum ventrale, pars caudalis –RA - robust nucleus of the aarchstriatum –area X the process of masculinization is dependent upon estrogen (aromatization of testosterone to estrogen)

28. Vocal Courtship in Songbirds Neurocircuitry: Neurocircuit involved in the production of learned song: HVcRAnXIIts muscles controlling syrinx production of sounds nXIIts: tracheosyringealis portion of the hypoglossal nucleus syrinx: vocal production organ in birds

29. Vocal Courtship in Songbirds Neurocircuitry: Neurocircuit involved in the acquisition of song –area X and MAN are important for learning song –hearing is important for learning song HVcRAnXIIts muscles controlling syrinx production of sounds auditory input area X thalamus MAN

30. Vocal Courtship in Songbirds Neurocircuitry: HVc is important for the production of learned song –lesion HVc will inhibit production of learned song (simple sounds remain) –lesions of RA or nXIIt will inhibit production of all sounds Area X and MAN are important for acquiring or learning song –if these areas are lesioned before crystallized song has been acquired, no crystallized song will occur –if these areas are lesioned after crystallized song has been acquired, crystallized song will occur; note that this would be true only during the current breeding season of seasonal breeders

31. Vocal Courtship in Songbirds Gonadal steroids affect this neurocircuitry in distinct ways: HVc, RA, MAN and area X are larger in males than in females HVc, RA and MAN all concentrate gonadal steroids –gonadal steroids act directly upon neurons within these three areas area X does not appear to concentrate gonadal steroids –it appears that neuronal input into area X from HVc is involved in masculinizing area X –survival or increased production of neurons in HVc leads to increased synaptic contact between neurons in these two areas –neuron--neuron interactions provide a trophic effect (increasing cell survival)

33. Vocal Courtship in Songbirds Seasonal changes in gonadal steroids and the display of song behavior and mating are reflected in seasonal changes in brain structure and connections. Ex. male canary: in this species, the male produces all or the majority of song male is an “open-ended learner”: –male goes through song learning and production each year –male learns new songs (e.g., adds song syllables to repertoire) seasonal variations in levels of testosterone and its metabolites: –during nonbreeding season, levels of testosterone are generally low; transient rises and falls in steroid levels are associated with increases in cell death followed by increases in neuronal recruitment within the brain; these changes reflect loss of some song syllables and the addition of others –during breeding season, levels of testosterone rise to peak values; high levels of testosterone are important for the production of crystallized song at high rates

34. Vocal Courtship in Songbirds males produce crystallized song males engage in mating A M JJ A SO N DJF M April June breeding season nonbreeding season High T levels Low T levels song production decreases form of song changes: –crystallized song changes to subsong or plastic song song learning occurs: –loss of song syllables –addition of new song syllables

35. Vocal Courtship in Songbirds How are seasonal changes in song production and learning mirrored by changes occurring within the brain? Study: Kirn et al. (1994) followed changes in the number of neurons within HVc during each month of one entire year (and after the first breeding season) in adult male canaries: –addition of neurons to HVc : administered tritiated thymidine into HVc of male birds during the beginning of each month and killed the birds 27 days later –death of neurons within HVc: identifying pycnotic profiles (dying neurons) discussed the relationship of these changes to seasonal changes that occur in learning new song syllables and in the levels of gonadal steroids

36. Vocal Courtship in Songbirds Neurocircuitry: Neurocircuit involved in the production of learned song: HVcRAnXIIts muscles controlling syrinx production of sounds nXIIts: tracheosyringealis portion of the hypoglossal nucleus syrinx: vocal production organ in birds

37. no change in neuronal recruitment or cell death A M JJ A SO N DJF M April June breeding season nonbreeding season Low T levels High T levels Low T levels Rise in T increase in pycnotic cells in HVc (cell death) increase in number of tritiated thymidine labeled cells in HVc (neuronal recruitment) increase in new song syllables Rise in T

38. Vocal Courtship in Songbirds Gonadal steroids could affect several processes leading to “neuronal recruitment” in HVc: –increased neurogenesis (more neurons are born) –increased migration –increased differentiation –increased survival A subsequent study suggests that gonadal steroids do not increase cell division (neurogenesis) but rather cell survival. neuroblast immature neuron migrate to new location mature neuron survive “neurogenesis” “differentiation”

39. Vocal Courtship in Songbirds Changes that occur in HVc leads to changes in other birdsong control regions. HVcRAnXIIts muscles controlling syrinx production of sounds Following a period of cell death, as new neurons are added to HVc, we see corresponding changes in the volume of RA: –increase in number of synapses –increase in dendritic length of neurons –increase in the size of neurons gonadal steroid receptors are present in both HVc and RA; the effects of testosterone (and/or its metabolites) on size of RA presumably reflects both direct actions on RA neurons and indirect actions via connections with HVc

40. Vocal Courtship in Songbirds Gonadal steroid-mediated plasticity can also be seen in the adult female canary. adult female canaries administer exogenous androgens activation of song behavior increase in size of HVc –(90% of male) –more neurons increase in size of RA –(50% of male) –larger neurons with longer dendrites and more synapses Complete masculinization of brain and song behavior is thought to be dependent upon gonadal steroids present during development and in the adult: –although exogenous androgens can activate song behavior in adult female canaries, the song that is produced by females is simpler in form –neuroanatomical changes that occur with androgen treatment in adult females do not reach the levels observed in normal adult males

41. Vocal Courtship in Songbirds Bottom line for role of gonadal steroids in song behavior by male songbirds: aromatization of testosterone to estrogen is important for masculinization of the songbird brain –HVc, RA, area X and MAN are larger in males than females estrogen is also important for acquisition of song (subsong and plastic song) testosterone is important for the crystallization of song, and for the activation of singing (courtship behavior) –these events occur seasonally in seasonal breeders testosterone (and/or its metabolites) is important for the addition of neurons in HVc, and for the formation of new connections with RA –these neuroanatomical changes are believed to reflect alterations in the bird’s song repertoire –these changes can be seen yearly in males that are seasonal breeders

42. Vocal Courtship in Songbirds Comparative studies of sex differences in song behavior: degree of sex differences in song behavior are mirrored by differences in brain Zebra finch Bay Wren only males singmales and females both sing (possess similar song repertoires) volume of HVc: males > females volume of HVc: males = females number of steroid- accumulating cells: males > females number of steroid- accumulating cells: males = females