Chp 4: Hormonal Influences on Male Sex Behavior

Male sex behavior can be divided into two main components:

• precopulatory behavior:

– production of ultrasonic vocalizations

– chemoinvestigation (sniffing of the female’s anogenital region)

• copulatory behavior:

– mounts: male mounts the female from behind

– intromissions: male, with an erected penis, inserts penis into the female’s

vagina

– ejaculations: explusion of a copulatory plug--composite of different

secretions from prostate gland, seminal vesicle and coagulating gland plus

sperm; this composite will “coagulate” forming a plug that serves to keep

the sperm within female’s reproductive system and increase likelihood of

fertilization

Sequence of Precopulatory and Copulatory Events:

chemoinvestigation

copulation

mount intromission ejaculation

(repeated X times)

postejaculatoryinterval(PEI)

satiety

postejaculatory interval (PEI): refractory period following ejaculation in which male does not engage in sex behavior (seconds to minutes)

satiety: refractory period following a series of ejaculations in which a male will not engage further in sex behavior with a given female

Quantification of Precopulatory & Copulatory Behaviors:

How to analyze male sex behavior?

• amount of time engaged in sniffing odors (chemoinvestigation)

• latency to mount; number of mounts

• latency to intromit; number of intromissions;

– interintromission interval--length of time between intromissions

• latency to ejaculate; number of ejaculations

• postejaculatory interval: length of time from an ejaculation to the next

intromission

Sexual MotivationIn addition to precopulatory and copulatory behaviors, we we can also consider

the male’s desire to engage in sex behavior.

• Sexual motivation: male’s inclination to seek out and approach a female for

the purpose of mating.

Tests of sexual motivation:

• latency to mount:

– mount quickly-->really motivated

– mount slowly-->not very motivated

– has problems--??

• Train a male to press a bar to gain access to a female--how many bar presses

will a male make? (more bar presses=greater motivation)

• Separate males and females across an electrified grid--how much electrical

current will a male withstand to gain access to a female? (greater current--

greater motivation)

Hormonal Influences on Male Sex Behavior

• In females, transient elevations of estrogen and/or progesterone during the

estrous cycle play an important role in activating female sex behavior.

• In males, testosterone (and/or its metabolites) also play an important role in

activating male sex behavior.

– Testosterone secretion increases at puberty: development of secondary sex

characteristics, increased muscle mass, production of sperm, and development of

an interest in sexual activity.

– However, testosterone levels do not show similar cyclical changes in levels

reminiscent of the female estrus cycle.

Hormonal Influences on Male Sex Behavior

Testosterone is the major androgen secreted by the testes.

• aromatization of testosterone to estrogen:

– important for activating male sex behavior in the rat (and in many other species,

although not all)

– involve aromatase and ERs

• reduction of testosterone to dihydrotestosterone:

– important for penile erections and intromissions in the rat

– involve 5-reductase and ARs

• testosterone as testosterone::

– important for aspects of sexual motivation

– involve ARs

Testosterone Secretion in Males:

• GnRH neurons secrete GnRH into the median eminence:

– GnRH stimulates release of LH and FSH from anterior pituitary

• In interstitial space (testis):– LH acts at the Leydig cells to

secrete the testosterone (“steroidogenesis”)

– LH-->increase levels of cAMP

• In seminiferous tubules:– at puberty, FSH and testosterone

act at Sertoli cells to initiate the production of sperm (“spermatogenesis”)

– FSH -->increase levels of cAMP

– FSH also stimulates the production of inhibin from Sertoli cells which inhibits release of FSH from anterior pituitary

GnRH Neuron

HYPO

ANTPIT

TESTES

FSHLH

Testosterone

GnRH

+

GnRH: gonadotropin-releasing hormoneFSH: follicle stimulating hormoneLH: luteinizing hormone

-

Inhibin

-

Hormonal Influences on Male Sex Behavior

Testosterone is the major androgen secreted by the testes.

• aromatization of testosterone to estrogen:

– important for activating male sex behavior in the rat (and in many other species,

although not all)

– involve aromatase and ERs

• reduction of testosterone to dihydrotestosterone:

– important for penile erections and intromissions in the rat

– involve 5-reductase and ARs

• testosterone as testosterone::

– important for aspects of sexual motivation

– involve ARs

Species Differences: Male Sex Behavior

• In rats (and hamsters):

– estrogen is critical for activating copulation--mounts, intromissions and

ejaculations )

– dihydrotestosterone is critical for penile reflexes

• In some species (lizards, mice &guinea pigs):

– dihydrotestosterone can stimulate male sex behavior

• In ferrets:

– testosterone plays selective role in sexual motivation

• In primates:

– testosterone may also have a selective effect on sex behavior not mediated

by estrogen or dihydrotestosterone

Peripheral Mechanisms:Androgens are critical for penile erections and deposition of the copulatory plug:

• androgens stimulate the size and number of sensory receptors on the surface of the

glans penis

• dihydrotestosterone (DHT) activates penile erections when rats are restrained on

their backs

• penile erections and deposition of the copulatory plug are controlled (in part) by

motoneurons within spinal nucleus of the bulbocavernosus:

– SNB innervates penile musculature

– DHT acts to increase survival of penile musculature followed by survival of SNB

motoneurons (during development)

– in the adult, DHT also affects the morphology of SNB neurons--increasing size of cell

bodies and increasing length of dendrities

– Note: penile erections and ejaculation are also controlled by the autonomic nervous

system.

Central Mechanisms:In the male rat:

• estrogen acts to stimulate copulation--mounts, intromissions and ejaculations

– aromatization is important for this process!

• MPOA plays an important role the effects of estrogen:

– sexually dimorphic area of the preoptic area (SDA-POA): sexual learning

– dorsal MPOA: critical for stimulation of mounts, intromissions and ejaculations

– dopaminergic inputs--activating mounts, intromissions and ejaculations

• amygdala plays an important role in facilitating sex behavior--the amygdala

attributes “salience” to cues associated with sex behavior

– corticomedial amygdala--pheromones to activate behavior

– basolateral amygdala--learned responses (e.g., bar pressing to gain access to female)

• dopaminergic inputs into the nucleus accumbens--sexual motivation

• dopaminergic inputs into the striatum--general locomotion (e.g., chasing female)

MPOAIn the rat:

• Aromatization of testosterone to estrogen is important for sexual differentiation of

the brain and activation of male sex behavior.

• The MPOA is an integrative center for hormonal stimulation of male copulation.

– Large lesions of the MPOA eliminate mounting, intromissions and ejaculations.

– Electrical stimulation of the MPOA stimulates these responses.

– Castrate an adult male rat and he will stop engaging in copulation.

– If you administer testosterone to a castrated male, he will again engage in copulation.

– If you administer testosterone or estrogen within the MPOA (with DHT to act

peripherally), normal male copulation will be seen-->MPOA is critical site!!

• The MPOA sends projections to neurons within the midbrain and brainstem, which

subsequently project to neurons in the spinal cord; 2 effects take place:

– activation of autonomic centers that control process of erection and ejaculation

– activation of motoneurons that control mounting and pelvic thrusts

MPOAAnatomical Subdivisions:

• Study by Heimer and Larsson (1966/67):

– Large bilateral lesions of the MPOA eliminated mating.

• Study by Arendash & Gorski (1983):

– Is the SDN-POA the critical brain site for control of male copulation?

– No: in sexually experienced males lesions of the SDN-POA had no significant effect on

male copulation.

– Instead, bilateral lesions of the dorsal MPOA produced substantial, long-term decreases in

number of mounts, intromissions, and ejaculations compared to sham-lesioned control

males.

• Study by De Jonge et al. (1989):

– Revisited role of SDN-POA in male sex behavior--found an effect!

– In sexually naïve males, bilateral lesions of the SDN-POA resulted in an increased latency

to first mount, intromission and ejaculation (took longer to initiate copulation)

– However, this decrement in sexual performance disappeared after repeated testing.

Amygdala:Corticomedial Amygdala (CMA):

• CMA: plays an important role in linking pheromones to activation of male sex

behavior

– CMA receives olfactory stimulation from the vomeronasal organ and olfactory mucosa

(sensory structures responsd to odors; project to the olfactory bulbs).

– Pheromones are secreted by female that can stimulate chemoinvestigation and copulation.

– CMA projects to the MPOA.

– Lesions of the vomeronasal organ and olfactory mucosa or bilateral lesions of the CMA

can eliminate male sex behavior--effect is similar to bilateral lesions of MPOA.

OlfactoryBulbs

CMA MPOA brainstem spinal cord

“Copulatoryreflexes”

Amygdala:Basolateral Amygdala (BLa):

• BLa: implicated in associative learning that feeds into motivation circuits.

• You can train a rat to bar press to gain access to an estrus female.

– If you lesion the BLa, the male rat will not show bar pressing for the female.

– However, if you place the receptive female in the chamber with the male, he readily

copulates with her.

– It is thought that the association between bar pressing to gain access to a female rat (for

copulation) has been lost.

• The BLa, via connections within the nucleus accumbens, is thought to play a role in

sexual motivation--searching out a female (e.g., bar pressing) in order to mate.

Nucleus AccumbensThe nucleus accumbens (NAcc) has been linked to motivational processes.

• Mogensen (1980) suggested that the NAcc serves as the “limbic-motor interface”

– a system concerned with translating emotional (“salient”) stimuli into motor output

BLa NAccVentral

Pallidum

Motor System:

Limbic System:

MPOA

Prefrontal Cortex

Basal ganglia

MesencephalicLocomotor

Region

“associativelearning”

a) foraging for a female

b) bar pressing for a female

“Motor Responses”

Central Mechanisms:In the male rat:

• estrogen acts to stimulate copulation--mounts, intromissions and ejaculations

– aromatization is important for this process!

• MPOA plays an important role the effects of estrogen:

– sexually dimorphic area of the preoptic area (SDA-POA): sexual learning

– dorsal MPOA: critical for stimulation of mounts, intromissions and ejaculations

– dopaminergic inputs--activating mounts, intromissions and ejaculations

• amygdala plays an important role in facilitating sex behavior--the amygdala

attributes “salience” to cues associated with sex behavior

– corticomedial amygdala--pheromones to activate behavior

– basolateral amygdala--learned responses (e.g., bar pressing to gain access to female)

• dopaminergic inputs into the nucleus accumbens--sexual motivation

• dopaminergic inputs into the striatum--general locomotion (e.g., chasing female)

Dopaminergic Cell Groups:

DopamineCell Bodies General Effect

incertohypothalamicDA cell group

ventral tegmentalarea (VTA)

substantia nigra dorsal striatumsensorimotorcoordination(locomotion)

nucleus accumbens

MPOAmounts, intromissionsand ejaculations(copulatory reflexes)

display of motivatedbehavior

Dopamine

The levels of dopamine increase within these areas during mating:

• Study by Pfaus et al. (1990):

– in vivo microdialysis was used to follow levels of dopamine (and its metabolites) within

nucleus accumbens and dorsal striatum during copulation

• in vivo microdialysis involves measurement of molecules present in the extracelular space

through diffusion across a semipermeable membrane at the base of a probe implanted into brain

– nucleus accumbens: levels of dopamine increased significantly during placement of a

sexually experienced male in a sex chamber (with odors of prior matings present), with

additional increases observed with placement of a female behind a screen, and then when

the male was allowed to mate with female

– dorsal striatum: levels of dopamine increased significantly only during interaction

between the male and female (copulation)

– concluded: DA released in the nucleus accumbens (from VTA) is involved in motivation

processes (prior to, and in preparation of, the actual act of copulation); DA released in the

striatum is involved in actual physical act of copulation (sensorimotor coordination)

Dopamine

The levels of dopamine increase within these areas during mating:

• Studies by Hull et al. (1995, 1997):

– in vivo microdialysis was used to follow levels of dopamine (and its metabolites) within

MPOA during copulation

– MPOA: levels of dopamine increased significantly during precopulatory period in which a

female is placed behind a perforated barrier (male can see, hear and smell female but not

mate with her), and then further increases in dopamine occur when the male can interact

with the female

– concluded: DA released within MPOA is involved in sexual motivation (DA is increased

prior to, and in preparation of, the actual act of copulation), and copulatory responses (DA

increases further during the actual act of copulation)

Central Mechanisms--Dopamine

Data from lesions and dopamine release during behavior provide insight into functional

roles of various brain regions to sex behavior:

• dorsal striatum:

– lesions of dopamine neurons in substantia nigra produce Parkinsonism symptoms:

difficulty initiating movements, slowness of actions and tremor

– DA levels increase significantly during the act of copulation

– dorsal striatum is thought to play a role in sensorimotor coordination (pursuit of female)

• MPOA:

– lesions of MPOA block display of mounts, intromissions and ejaculations

– administration of DA antagonists into the MPOA can inhibit display of copulation

– DA levels increase prior to, and during, the act of copulating with a receptive female

– MPOA is thought to play a role in both sexual motivation and copulation

– MPOA receives pheromone-associated information from CMA; this information provides

one source of excitatory information linked to both sexual motivation and performance

Central Mechanisms--Dopamine

Nucleus Accumbens (NAcc):

– lesions do not block the display of copulation

– however, blocking input into the NAcc by bilateral lesioning the BLa can block bar

pressing in order to gain access to a receptive female (discussed in terms of sexual

motivation)

– DA levels increase prior to, and during, the act of copulating with a receptive female

– however, DA levels also increase in NAcc in response to feeding, drinking, drug self-

administration

– Nacc is thought to play a role motivational processes and reward

Hormonal Regulation

Testosterone, and/or its conversion to estrogen, is important not only for stimulating

copulation but also for motivational processes.

If you train a male to bar press to gain access to a female, you can study the effects of

specific brain lesions and compare those effects to that produced by castration.

Sexual performance:

• bilateral lesions of the MPOA, but not BLa, eliminate copulation (low % of rats

engaged in copulation)

• castration also dramatically decreases % rats copulating

Motivation processes:

• bilateral lesions of the BLa, but not the MPOA, greatly reduce bar pressing to gain

access to a receptive female

• castration also dramatically decreases bar pressing

Hormones and Dopamine Regulation

Does castration alter dopaminergic neurotransmission to effect changes in male sex

behavior? Answer--yes, testosterone facilitates dopamine release!

Studies by Alderson & Baum (1981); Mitchell & Stewart (1989):

• castration decreases the level of dopamine within the nucleus accumbens

• administration of testosterone, or estrogen plus dihydrotestosterone, increases

dopamine levels within this nucleus

Study by Hull et al. (1997):

• castration decreases release of dopamine within the MPOA in response to a

precopulatory period (female behind barrier) or interactions with an estrous female

• some castrates (1 week) showed dopamine release in MPOA and engaged in sex

behavior (short-term response; eventually all castrated males will stop mating)

• administration of testosterone proprionate restored release of dopamine within

MPOA in response to precopulatory period and facilitated mating

Opioid and Male Sex Behavior

Opioids have been shown to inhibit male sex behavior.

Study by Hughes & colleagues (1988):

• infusion of B-endorphin into the medial preoptic area causes a reduction in male’s

mating performance

• infusion of naloxone, an opioid receptor antagonist, can facilitate sexual

performance and block inhibitory effect of B-endorphin

MPOA ArcuateNucleus

B-endorphinneurons

malecopulation

naloxone

Opioid and Male Sex BehaviorOpioids have been shown to stimulate male sex behavior.

Mehrara and Baum (1990):

• Design:

– conditioned place preference is a test of motivation (desire to spend more time on one side of a chamber than the other)

– established conditioned place preference in male rats given the opportunity to mate on only one side of the chamber

– how much time would the males spend on the side of the chamber in which they had mated

– several variables: 1) sham-operated vs. castration, 2) given naloxone at varying doses or saline, and 3) testing the

animals 7 days after surgery or 14 days

Opioid and Male Sex Behavior

Opioids have been shown to stimulate male sex behavior.

Mehrara and Baum (1990):

• Results:

– administration of naloxone decreased time spent in the chamber associated with mating

(e.g., loss in conditioned place preference)

– administration of naloxone also decreased time spent in the chamber associated with

mating in males 7 days after castration, but not 14 days, after castration

• Conclusion:

– opioids act to facilitate conditioned place preference (blocking opioid activity can

blockthis response)

– the ability of opioids to facilitate conditioned place preference is lost at 14 days

postcastration

– opioids act to stimulate dopamine release from neurons within the VTA; dopamine

neurons in VTA project to NAcc; NAcc is involved in motivational processes

Opioid and Male Sex Behavior

Opioids have been shown to stimulate male sex behavior.

+

Nucleus Accumbens

opioids

dopamineneurons

Motivation(e.g., conditionedplace preference)

+VTA

Neurons & Behavior

How to study role of specific groups of neurons in specific behavioral responses?

• compare the behavior of lesioned animals to sham-operated controls

– lesions can be made with electrical current (electrolytic) or with neurotoxins

(neurochemical)

• compare the behavior of different groups of animals following the administration of

drugs that either increase or decrease the level of a specific neurotransmitter:

– lesion specific neurons: 6-hydroxydopamine is used to lesion dopamine neurons

– use agonists to stimulate activity at specific receptors

– use antagonists to block activity at specific receptors

• follow the activation of groups of neurons at various endpoints of behavior:

– release of neurotransmitters

– expression of molecules that serve to “mark” activated neurons

• expression of c-fos proto-oncogene

c-fos Expression

c-fos expression is used as a marker of neuronal activation:

• levels of c-fos mRNA and Fos protein are low in control, unhandled animals

• if you stimulate an animal--handling, mating, stress, etc., you will see significant

increases in the level of these molecules

• the rise and fall occurs quickly:

– c-fos mRNA peaks by 30-60 minutes, and declines to basal levels within 2 hours

– Fos protein peaks by 1-2 hour,s and declines to basal levels within 4 hours

• the transient nature of c-fos expression as well as the low levels basally enables

detection of these molecules to serve as a “marker” of neuronal activation

• c-fos expression serves to couple “inputs” (neuronal or hormonal) with gene

transcription:

– Fos protein forms a dimer with Jun protein--called “AP-1” factor

– AP-1 factor binds to specific DNA sequences (AP-1 site) to control transcription of genes

Components of Mating

Kollack-Walker & Newman (1997):

• Experimental Design:

– sexually experienced male hamsters were exposed to female hamster vaginal secretion

(FHVS) for 10 minutes, or allowed to mate to different endpoints of copulation: 5

intromissions, 1 ejaculation, 5 ejaculations, long intromissions

– long-intromissions: characterized by a period of extended intravaginal thrusting during

which no sperm transfer occurs; the occurrence of long intromissions is believed to reflect

a state of sexual satiety (turning off male sex behavior)

– following FHVS exposure or copulation, males were kept in the mating arena (up to one

hour) until they were anesthetized, perfused with fixative, their brains removed from the

skull and sectioned on a freezing microtome

– a series of sections from each animal were processed for Fos immunocytochemistry

– the number of Fos-immunoreactive neurons were determined for specific brain regions in

each animal, averaged per group, and compared statistically

Components of Mating

Kollack-Walker & Newman (1997)

• Results:

– significant patterns of activation were observed within medial amygdala (MeA), bed

nucleus of the stria terminalis (BNST) and medial preoptic area (MPOA) following:

FHVS exposure alone, mating to 1 ejaculation, and mating to multiple ejaculations

– in particular, neuronal cell “clusters” in the caudal part of the posterior medial nucleus of

the amygdala (cMePD) were seen in 67% of males mating to 5 ejaculations, and in 100%

of males mating to long intromissions

• Conclusion:

– different patterns of c-fos expression were observed in males that engaged in different

levels of sexual activity (chemoinvestigation, copulation)

• Question: Does the selective pattern of neuronal activation seen following multiple

ejaculations reflect a state of satiety or simply a phenomonon of experiencing

multiple ejaculations?

Sexual SatietyParfitt & Newman (1997)

• Experiment #1: Do the number of ejaculations prior to the occurrence of long

intromissions make a difference in the pattern of c-fos expression?

• Design:

– compare pattern of Fos-immunostaining within sexually experienced male hamsters mated to

long intromissions in 2 groups: 1) rested male hamsters (rested 1 week), and 2) male

hamsters mated to long intromissions on 4 consecutive days prior to experimental day

– rested male hamsters show more ejaculations (on average 9 ejaculations) while repeatedly

mated males show fewer ejaculations (on average 4 ejaculations)

• Results: (focus on neuronal cell “clusters” in cMePD)

– males mated to long intromissions in both groups (rested male hamsters and repeatedly

mated male hamsters) showed the presence of neuronal cell clusters in cMePD

• Conclusion:

– the number of ejaculations does not alter presence of neuronal cell“clusters” in cMePD

Sexual SatietyParfitt & Newman (1997)

• Experiment #2: Does mating to only 1 ejaculation produce different patterns of

neuronal activation in males that differ in their proximity to sexual satiety?

• Design:

– compare pattern of Fos-immunostaining within sexually experienced male hamsters mated

to only one ejaculation in 2 groups: 1) rested male hamsters (rested 1 week), and 2) male

hamsters mated to long intromissions on 4 consecutive days prior to experimental day

• Results: (focus on neuronal cell “clusters” in cMePD)

– following mating to only 1 ejaculation, repeatedly mated males showed the presence of

neuronal cell “clusters” within cMePD, while the rested males did not

• Conclusion:

– the number of ejaculations are not critical for the occurrence of neuronal cell “clusters”

within cMePD, rather the proximity to satiety was the critical factor

– neuronal cell “clusters in cMePD may reflect development of a state of satiety!

The Coolidge Effect

The Coolidge Effect:

• males recently mated with a particular female appear uninterested in further

copulation (sexual satiation)

• however, if a new female is introduced, then the male will continue to mate

• thus, the introduction of novel females can extend a male’s sexual motivation and

performance

Sexual Satiety

Summary:

• different patterns of c-fos expression are observed in male hamsters mated to

different endpoints of copulation--female odors, 1 ejaculation and multiple

ejaculations

• neuronal cell “clusters” in cMePD: their appearance is dependent upon how close a

male is to sexual satiety

– clusters are present in all males mated to long intromissions (LI)

– clusters are present after only 1 ejaculation in repeatedly mated males (4 ejaculations away

from LI) but not in rested males (9 ejaculations away from LI)

• thus, neuronal cell “clusters” in cMePD may reflect development of a state of satiety

• in the literature, there is evidence that lesioning this part of the medial amygdala in

female hamsters can produce a significant increase in the duration of copulation

[Takahashi & Gladstone (1988) Behav. Neurosci., 102: 268-275]

Seasonality in Reproduction

• Seasonal breeders: engage in sexual activity during certain times of the year.

• Reproductive axis also varies with the breeding season.

• Seasonal breeders: hamsters, sheep, deer, zebra finches, ferrets

• Nonseasonal breeders: rats, humans, primates

• Similar events within the reproductive axis are observed in males and females of

species that are seasonal breeders.

During Breeding Season Out of Breeding Season

GnRH

LH & FSH

testes

testosterone

GnRH

LH & FSH

testes

testosterone

Display of sex behavior No display of sex behavior

Seasonality in Reproduction

• The breeding season for different species varies, and is dependent upon an

interaction between environmental factors and length of gestation.

• Environmental factors: food availability, temperature--signaled by changes in day

length (photoperiod).

• Goal: ensure that the offspring are born under conditions optimal for their survival.

Ex. Sheep Ex. Ferrets

Breeding season in fall Breeding season in spring

Daylength is decreasing Daylength is increasing

Gestation period: 5 months Gestation period: 41 days

Newborn lambs will be bornin spring

Newborn ferrets will be bornin spring

Seasonality in Reproduction

• Seasonality can affect how the brain responds to gonadal steroids:

• How well does administration of exogenous testosterone restore male sex

behavior?

• Testosterone is most effective in stimulating male sex behavior in males housed

under long day photoperiod compared to males housed under short day

photoperiod.

malehamsters

castrate replacetestosterone

house undershort day

photoperiod

house underlong day

photoperiod