Outline of today’s lecture

2. Outline of today’s lecture • Part I • Why YOU should care about hormones? • Definition--What is a hormone? • Introduction to behavioral endocrinology (4 levels of analysis) • Common techniques in behavioral endocrinology (50% of today) • Part II • The endocrine system (other 50%) • major hormones: hypothalamic, pituitary, thyroid, GI, pancreatic, steroid, monoamines • Regulation • Revisiting question #1 and comparison with fMRI

3. Why should social-personality psychologists study hormones? • Conservative estimate--the human brain is riddled with billions and billions of endocrine receptors? • Prima facie evidence of hormonal influence on behavior, thought, mood, emotion, personality? • Ignorance of the hormone-behavior link could have dire consequences • Corticotropin-releasing hormone (CRH) is a key neuroendocrine factor implementing endocrine, immune and behavioral responses to stress. The expression of CRH receptors was analyzed for the first time in pituitaries of suicide victims by in situ hybridization (2001--Molec. Psych). There was a shift in the ratio of the two major CRH receptors (R1 and R2) in the pituitaries of suicide victims, relative to those who died of natural causes. Causality unclear. • Today’s lecture does NOT focus on the hormone-behavior link. That stuff you can pick up from journal articles, with new and exciting research coming out each week! Today we need to cover the bedrock, the core, the basics, the stuff that you need to function effectively and intelligently in this world. • And just to keep you from drifting off during the biological onslaught that is about to hit, let’s take a peek at the final slide next.

4. Testosterone plays a crucial role in neuronal function, but elevated concentrations may have deleterious effects. Here it is shown that supraphysiological levels of testosterone (micromolar range) initiate the apoptotic cascade. Short periods of elevated testosterone levels (six to 12 hours), such as those resulting from the use of muscle-building steroids,may lead to "cell death" and may have long term effects on brain function. What is a hormone? • Hormones coordinate the physiology and behavior of an animal by regulating, integrating, and controlling its bodily functions. • Example: The same hormone (e.g., Luteinzing Hormone--LH) that causes egg or sperm maturation also stimulates mating behavior in many species. • This dual function ensures that mating occurs ONLY when animals have mature gametes (eggs or sperm) available for fertilization. • Hormones are similar to neurotransmitters, but can operate over a greater distance and over a much greater temporal range than neurotransmitters. • Differences between hormones and neurotransmitters: • Neural messages can only travel along existing nerve tracts; hormonal messages can travel in the circulatory system; thus any cell receiving blood is potentially able to receive a message. • Neural messages are digital, all-or-none events that have rapid onset and offset; neural signals can take place in milliseconds; plus, electrical signal can travel along myelinated axons at speeds up to 100 meters per sec! Hormonal messages are analog, graded events that can take seconds, minutes or hours to occur (more detail to follow). • How does a hormone exert its influence? • Only cells with receptors for that hormone can be influenced • Called target cells • Interaction of a hormone with its receptor leads to a genomic response whereby the hormone activates genes that regulate protein synthesis (e.g., up-regulation: synthesis of a receptor for that hormone). • Some hormone effects are nongenomic.The monoamines. • Nongenomic (transcription-independent) effects are principally characterized by their insensitivity toinhibitors of transcription and protein synthesis. The most obvious experimental evidence suggesting their existence is rapid onset ofaction (within seconds to minutes). These rapid effects are likely notbe mediated through intracellular receptors. The dual effect of LH: LH stimulates gonads to produce gametes, and stimulates gonads to produce testosterone Action potentials propagate faster in axons of larger diameter, other things being equal. They typically travel from 10-100 m/s.

5. Hormonal Effects Androgen receptor (computer image, left; electron micrograph, right) • Sufficient number of receptors must be available for hormonal effects to occur. • Popular belief that individual differences in behavior reflects differences in hormone concentrations. For example, it is assumed that roosters that crow frequently have more testosterone than roosters that seldom crow (or that aggressive men have higher T). • Not necessarily true! • Individual differences in behavior can reflect hormone concentrations,pattern of hormone release, numbers and location of hormone receptors, and the efficiency of those receptors in affecting gene transcription. • Hormones rarely change the function of a cell; rather, they alter the rate of normal cellular function. • Thus, hormones affect cell morphology and size (including development of muscle and neuronal cells), and affect cell death (apoptosis) throughout the nervous system. • Although hormones obviously affect behavior, it is also true that behavior can influence hormonal levels and hormonal effects.

6. How might behavior affect hormones (most research does not look at this) • Behavior can and often does affect hormone levels which in turn can influence subsequent behavior. • World Cup Soccer Fans were assayed for testosterone before and after the Brazil-Italy final. Brazil won on penalty kicks. 11/12 Brazil fans showed an increase in testosterone, whereas 9 of 9 Italian fans showed a decrease. • Testosterone concentrations were measured in four heterosexual couples over a total of 22 evenings. On 11 evenings, saliva samples were obtained before and after sex; on the remaining 11 evening, two samples were obtained, but there was no sex. Having sex caused an increase in testosterone in both men and women. No changes were seen in the no-sex nights. The early evening samples revealed no difference in testosterone concentrations between sex and no-sex evenings, suggesting that sex increases testosterone more than testosterone (concentrations) cause sex. Alternatively, physical exercise may have caused the increase (it increases CORT, which can correlate positively with T). little Dutch football fan

7. How does one go about answering a research question in the field of behavioral endocrinology? Example: What causes the Zebra Finch to sing? • (What causes Zebra Finches to Sing?) • Four correct answers, based on Levels of Analysis • Immediate causation: mechanisms mediated by the nervous and endocrine systems • Example: Singing in male zebra finches (In contrast to mammals in which structural differences in neural tissues have not been directly linked to behavior, structural differences in avian brains have been directly linked to a sexually dimorphic behavior: bird song). • Female zebra finches never sing, even after testosterone treatment in adulthood. • Other species (wrens, canaries) show no or a diminished sex difference • The size of nuclei in two major brain circuits (HVc, RA, & Area X) implicated in learning and production of bird song parallel sex differences in singing behavior (e.g., large dimorphisms in zebra finches,undetectable dimorphisms in wrens in which no singing difference is observed). • But why the sex difference in the zebra finch? • If female finches are treated with androgen soon after hatching, and then treated with androgen when adults, they don’t sing, but they showed a small increase in the number of neurons in the song production region. • If female finches are treated with estrogen soon after hatching, then as adults, they show a marked increase in the number and size of neurons in this region, but still no singing. • However, if treating with estrogen soon after hatching and then treated with androgen as adults, they show the same size and number of neurons as their male conspecifics, and they SING. Conclusion: Estrogens are necessary to organize the neural machinery underlying the song system, and androgens activate it. Bird testes produce circulating androgens which enter neurons containing aromatse, an enzyme which converts androgens to estrogen. These neurons are generally found in the hypothalamus, as well as in the structures constituting the neural circuit controlling bird song. • Development Behavioral responses change through the lifespan as a result of gene X environment interaction. The mating dance of columba chippendalia is virtually unique in the animal kingdom

8. Levels of Analysis (Zebra Finch song) • Hormonal events affecting the fetal and neonate can have profound consequences later in life. • Most research has focused on how early events influence adult behavior; however, the decay of behavioral patterns during aging is also a new and expanding area to those pursuing developmental questions. • Possibly, zebra finches sing because they have undergone puberty or because they learned songs from their fathers. • Evolution • This approach involves many generations of animals and addresses the ways that specific behaviors change during the course of natural selection. • Biologists study the evolutionary bases of behavior in order to learn why behavior varies between closely related species as well as to understand the specific behavioral changes that occur during the evolution of a new species. • Behaviors rarely leave interpretable traces in the fossil record, so this approach relies upon comparing existing species that vary in relatedness (e.g., old v. new world monkeys). • Someone at this level might say that zebra finches sing because they are finches, and that all finches sing because they have evolved from a common ancestral species that sang. • Adaptive Function • Synonymous with adaptive significance; role that behavior plays in the adaptation of animals to their environment and with the selective forces that currently maintain behavior. • Could be argued that male zebra finches sing because it will increase the likelihood of reproduction by attracting females to their territories and dissuading competing males from entering. • So, if we want to study HOW, then focus on questions of immediate causation and development. • If WHY, then questions of evolution and function.

9. How might hormones affect behavior • The study of the hormone-behavior relationship is organized around the idea that animals are composed of three interacting components: • Input systems (sensory) • Integrators (CNS) • Output systems (e.g., muscles) • Example: Removing the testes of the male zebra finch stops it from singing. Reinplant the testes, or provide the primary testicular hormone, testosterone, and singing resumes. Obviously, testosterone is involved in singing, but how? • Input: Examine the sensory system. Does testosterone alter the birds’ sensory capabilities, making the environmental cues that elicit singing more salient? If this were the case, females or intruders might be seen or heard more easily. • CNS: Testosterone could change the Neural Architecture or speed of neural processing. Higher processes (e.g., motivation, attention) might be influenced. • Effectors: Testosterone concentrations might affect the muscles of the syrinx (the avian vocal organ). • This 3-part framework can aid in the design of hypothesis and experiments to help understand how hormones affect behavior.

10. Classes of evidence for determining hormone-behavior interactions • 1st: A hormonally-dependent behavior should disappear when the hormonal source is removed or actions of the hormone are blocked. Example--ADT. • 2nd: After the behavior stops, restoration of the missing source or its hormone should reinstate the absent behavior. Again, ADT. • 3rd: Hormone concentrations and the behavior should covary; in practice, the behavior should be observed when concentrations are relatively high and never or rarely observed when concentrations are low. • This 3rd class of evidence is difficult to obtain because many hormones have a long latency of action (why? up-regulation.)and/or a long offset latency (why? down-regulation) and are released in a pulsatile manner. For example, a pulse may be released into the blood and then no more released for an hour or more, so a single sample will not provide an accurate picture of the endocrine status of the animal. • Another problem is that biologically effective amounts of hormones are TINY and thus difficult to measure accurately. Effective concentrations are measured in micrograms, nanograms, or picograms (10 to the negative 6th, 9th, or 12th, respectively). • Unfortunately, the 1st two classes of evidence are thus considered more reliable, but research on humans is typically limited to the 3rd (with exceptions--ADT, for example).

11. Common techniques in behavioral endocrinology • Ablation & replacement • Bioassays • Immunoassays • Immunocytochemistry (ICC) • Autoradiography • Blot tests • In situ hybridization • Pharmacological Techniques • Genetic Techniques (transgenics and knockouts)

12. Common techniques in behavioral endocrinology • Ablation and replacement • 1. A gland that is suspected to the the source of the hormone affecting behavior is surgically removed • 2. Effects on behavior are observed • 3. Hormone is replaced, by reinplantation, injection of an extract from the gland or injecting a purified hormone • 4. Determination is made whether the observed consequences of ablation are reversed by replacement therapy. • 5. Is the surgically removed gland the hormonal source?

13. Common techniques in behavioral endocrinology • Bioassays Once the existence of a hormone has been established, the next step is to identify the chemical processes involved in its actions. • Typically, this involved a test of its effects on a living animal (which can serve as a reliable, quantifiable response system on which to test hormonal extracts and chemical fractions). • A bioassay need not be conducted on the same species from which the hormone was obtained. • Example: THE RABBIT TEST (or Friedman test). • Developed by Maurice Friedman in 1929 (used until the late 1950s). Used to test for the presence of human chorionic gonadotropic (hCG--a hormone released from the implantation site of a blastocyst. hCG prevents menstruation). hCG found in women’s urine. • BTW, hCG produced by the rudimentary placenta that forms immediately after blastocyst formation (hCG maintains corpus luteal function during pregnancy--thus, progesterone secretion--and inhibits ovulation). • Urine injected into a rabbit, and if hCG present, rabbit’s ovaries would form corpora lutea, or “yellow bodies” (temporary ovarian endocrine structures formed following ovulation within 48 hours and produce progestins--horomones that support pregnancy). • It is a common misconception that the injected rabbit would die only if the woman was pregnant. This led to the phrase "the rabbit died" being used as a euphemism for a positive pregnancy test. In fact, all rabbits used for the test died, because they had to be surgically opened in order to examine the ovaries. While it was possible to do this without killing the rabbit, it was generally deemed not worth the trouble and expense. • The rabbit test was better than the earlier mouse test (developed in 1928) which required 6 or more mice and 96 hours to complete. • However, if the rabbit was stressed, spontaneous corpora lutea formation occurs (in the absence of hCG), so there was a significant false positive rate. • In the frog test for pregnancy, a woman’s urine is injected into a male frog or toad. In 2-4 hours, the animal would begin to produce sperm if the woman’s urine contained hCG. However, there were seasonal variations in frogs--in the summer, they had a greater tendency to produce false negatives. corpus luteum

14. Common techniques in behavioral endocrinology • Immunoassays Bioassays require a great deal of time, labor, and the sacrifice of many animals for every assay. • The development of the radioimmunoassay (RIA) reduced these problems and increase the precision with which hormone concentrations could be measured. • Based on competitive binding of an antibody to its antigen. An antibody produced in response to any antigen (defn: any molecule that stimulates an immune response) has a binding site THAT IS SPECIFIC FOR THAT ANTIGEN. • Antigen molecules can be labeled with radioactivity, and an antibody cannot discriminate between an antigen that has been radiolabeled (or “hot”) and one that is normal (or “cold”). A given amount of antibody possesses a given number of binding sites for its antigen. STEPS INVOLVED IN RIA: • 1. First, inject the hormone of interest (e.g., T) into an animal to raise antibody (anti-T) • 2. Then, collect antibody from blood, and purify. • 3. Develop a standard curve • Set up 5 or 6 reaction tubes, each containing the same amount of antibody, the same amount of radiolabeled hormone, and different amounts of cold purified hormone of known concentrations (from low to high concentration) . • (The radiolabeled and cold hormone compete for binding sites on the antibody, so the more cold hormone present, the less hot hormone will bind to the antibody) • 4. The quantity of hot hormone that was bound can be determined by precipitating the antibody and measuring its radioactivity. • 5. The concentration of hormone in an UNKNOWN sample can then be determined by subjecting it to the same procedure (substituting unknown sample for cold hormone in STEP 3) and comparing the results with the standard curve • The enzymoimmunoassay (EIA) works on the principle of competitive binding of an antibody to its antigen. The major difference is that EIAs do not require radioactive tags. Rather, the antibody is tagged with a compound that changes optical density (color) in response to binding with the antigen. • Example: The home pregnancy test. • However, most EIAs provided quantitative information, and thus a standard curve is generated, so that different amounts of the hormone in question provide a color gradient that is read on a spectromoter. A similar technique is called enzyme-linked immunosorbent assay (ELISA).

15. Animation of an HIV ELISA test (both positive and negative test--notice that you can test for presence of antigen (e.g., a hormone assay) or its complement antibody (viral assay). http://www.biology.arizona.edu/IMMUNOLOGY/activities/elisa/technique.html Anatomy of an ELISA test

16. Common techniques in behavioral endocrinology • Immunocytochemistry (ICC) • ICC techniques use antibodies to determine the location of a hormone in the body. • Antibody molecules linked to marker molecules (usually a fluorescent dye) are introduced into dissected tissue from an animal, where they bind with the hormone or neurotransmitter of interest. • Tissue is examined under a fluorescent microscope, and concentrated spots of fluorescence will appear, indicating where the protein hormone is located. • Commonly used marker is the enzyme horseradish peroxidase. human sputum cells

17. Common techniques in behavioral endocrinology • Autoradiography • Typically used to determine hormonal uptake and indicate receptor location. • An animal can be injected with a radiolabeled hormone, or the study can be conducted in vitro. • Top picture: Human NMDA receptor (NDMA is a receptor for the amino acid glutamate, which is the most abundant neurotransmitter in the mammalian nervous system). • Bottom indicates transport in a young tomato plant: The distribution of elements (e.g. micronutrients, pollutants) are visualized by autoradiographic techniques.

18. Other techniques in behavioral endocrinology • Blot tests (uses a technique called gel electrophoresis to separate proteins based on their length and weight (in kDa) • Used to determine presence of a particular protein or nucleic acid in a specific tissue. • Southern blot used to assay DNA • Northern blot used to assay RNA • Western blot used to assay proteins • In Situ Hybridization • Previous techniques can determine only whether or not a particular substance is present in a specific tissue, but in situ hybridization can be used to determine if the substance is produced in a specific tissue. • Used at the cellular level to examine gene expression. • More specifically, used to identify cells that are producing mRNA for a specific protein (e.g., a hormone or neurotransmitter). • Called hybridization because a radiolabeled cDNA probe (cDNA, or complementary DNA is synthesized from mature mRNA by the enzyme reverse transriptase) is introduced into the tissue. If the mRNA of interest is present, the cDNA will form a tight association (i.e., hybridize) with it. • These are chromosomes from the canola seed showing the location of various retrotransposons, which are genetic elements that can amplify themselves within a genome.

19. Other techniques in behavioral endocrinology • Pharmacological Techniques • Use of synthetic agonists (mimics) and antagonists (blockers) to determine endocrine functioning. • Some agents act to stimulate or inhibit endocrine functioning by affecting the release of hormones; they are called general agonists/antagonists. • Others act directly on receptors, enhancing or negating the effects of the focal hormone; these are receptor agonists/antagonists. • Example: CPA is a powerful anti-androgen used clinically to treat male sex offenders (about 20% of patient don’t show the expected behavioral response). CPA binds to androgen receptors but doesn’t activate them, thereby blocking effects of androgen.

20. Other techniques in behavioral endocrinology • Common genetic manipulations in behavioral endocrinology are the insertion (transgenic) or removal (knockout) of the genetic instructions encoding a hormone or hormone receptor. • Briefly: The genetic instructions for each individual are contained in the DNA, located in the nucleus of nearly every cell. • Each gene (composed of a specific order of four nucleotides: adenine, thymine, cytosine, and guanine) is determined by the sequence of nucleotides along the “rails” of the double helix. • To inactivate (knockout) a gene, you scramble the order of the nucleotides that make up the gene. The identification of the genome has been most successful in the mouse; thus mice are most commonly used in knockout studies. • Knocking out a gene is more difficult than it sounds: • gene of interest must be identified, targeted, and marked precisely. • A mutated form of the gene is then created (e.g., mutating the marker gene via genetic engineering). • Embryonic stem cells are harvested and cultured, and the mutated gene is introduced into the cultured cells by microinjection. • A small number of the altered genes are incorporated into the DNA of the stem cells via recombination. • The mutated embryonic stem cells are inserted into normal embryos (blastocycts), which are then implanted into surrogate mothers. • That’s it!!!! • All of the cells from the mutated stem cells will have the altered gene; the descendents of the normal embryonic cells will have normal genes. • Thus, the offspring will have a mixture of cells--some containing the mutated gene and some containing the normal (wild-type) gene. • This animal is called a chimera • The chimeras are then bred (interbred or bred with wild-type animals) to produce wild-type (+/+), heterozygous (+/-), and homozygous (-/-) animals with respect to that gene. Behavioral performance can then be compared. n.b In Greek mythology, a Chimera is a monster, depicted as an animal with the head of a lion, the body of a she-goat, and the tail of a dragon

21. Chimeras • Chimeric animals: • Pictured on the right is a baby “geep”, made by combining a goat and sheep embryo. Notice the chimerism evident in the skin - big patches of skin on front and rear legs are covered with wool, representing the sheep contribution of the animal, while a majority of the remainder of the body is covered with hair, being derived from goat cells. • There is also some potential that this technique can be applied to problems such as rescue of endangered species. It is possible, for example to construct a goat-sheep chimera such that a goat fetus is "encased" in a sheep placenta. This enables a sheep to carry a goat to term, which will not occur if you simply transfer goat embryos into sheep (the sheep will immunologically reject the goat placenta and fetus). It may be possible to extend this procedure to allow embryos from severely endangered species to be carried by recipient mothers from another species.

22. Fat mice--Using all of the techniques discussed previously to understand obesity • Effects of Leptin • Recently leptin (derived from Greek leptos, meaning thin) discovered as a hormone released from fat cells. • It is known that a specific mutation in mice on the ob gene can cause extreme obesity (in mice homozygous for defective ob). • Thus, these naturally-occuring mutants can be considered “natural knockouts” for the ob gene. • ob/ob mice have a pair of defective ob genes, overeat, are obese, and are sterile. • The ob gene normally codes for leptin, which is released into the bloodstream and travels to specific receptors in the CNS and elsewhere to regulate feeding and energy balance. • Although it has been known for a long time that the ob/ob mutationaffects body weight in mice, only recently has the ob gene been cloned, inserted into a bacterial system, and thus purified leptin made available to researchers. • Replacement studies could now be done in which leptin was provided to ob/ob mice to determine if leptin replacement would ameliorate obesity. • It did (see mouse on the right (25g--average mouse is 15g; one on the left is w/out replacement at 35g). • Availability of purified leptin allowed for the development of antibodies used to develop assays to determine blood concentrations. • RIA determined that there was no connection between plasma leptin concentrations and obesity/diatetes IN HUMANS. • A leptin ELISA was developed for rats, which determined that fasting or exposure to low temperatures caused leptin to fall. • Immunocytochemistry determined that leptin was present in both white (energy utilization) and brown (generation of body heat) adipose tissue. • Autoradiography determined that tagged leptin was found in a brain area located in the front of the third ventricle. • This tissue was then used to clone a leptin receptor • In situ hybridization found that the mRNA for the leptin receptor was expressed in the hypothalamus. • Efforts to “cure” obesity involved transgenics--treating ob/ob mice with a recombinant virus expressing mouse leptin cDNA, resulting in leptin production. A dramatic reduction in food intake and body mass resulted. • This treatment also reverses the sterility found in ob/ob mice. • However, only two (out of thousands) of obese humans displayed a ob mutation. Sadly, the effects of leptin on humans have been disappointing, dashing the Nobel dreams of more than a handful of psychologists.

23. The Endocrine System • Where do hormones come from? • They are produced by glands, and are secreted into the bloodstream. • Where do hormones go? • They travel to target tissues containing hormone-specific receptors. • What do hormones do? • By interacting with their receptors, they initiate biochemical events that activate genes to induce certain biological responses (e.g., protein synthesis). In some cases, hormone-receptor interactions result in nongenomic effects on cellular function (these are fast, and are just now being studied in their role in mediating behavior).

24. Types of chemical communication • Intracrine mediation • regulation of intracellular events • Autocrine mediation • autocrine substances feedback to influence the same cells that secreted them. • Paracrine mediation • paracrine cells secrete substances that affect adjacent cells (e.g., nerve cells). • Endocrine mediation • endocrine cells secrete chemicals into the bloodstream where they may travel to distant target cells. • Ectocrine mediation • Ectocrine substances, such as pheromones, are released into the environment to communicate with others.

25. General Features of the Endocrine System • Endocrine glands are ductless • Endocrine glands have a rich blood supply. • Product of endocrine glands (hormones) are secreted into the blood stream • Hormones can travel to virtually any cell in the body (cells in the lenses of the eye are an exception--no blood supply) • Hormone receptors are specific binding sites, embedded in the cell membrane (in the case of peptide hormones) or in the cytoplasm (in the case of steroid hormones) that interact with a hormone or class of hormones • Exocrine glands have ducts or tubes (e.g., salivary, sweat, mammary). Some glands have both endo- and exo- structures (e.g., pancreas)

26. The Endocrine Glands In touch with his feminine side

27. Four classes of hormones Protein & peptide hormones Steroid hormones Monoamines Lipid-based hormones (prostaglandins)

28. Cellular and molecular mechanisms of hormone action--Protein Hormones (the majority of hormones) • Protein hormones require a second messenger (i.e., a molecular middleman--typically, an enzyme or another protein) to transduce (conversion of one type of signal into another) the hormonal signal • In really simple terms, the hormone binds to the receptor which is coupled to a protein called G. When this mess is formed, a messenger called cAMP is created. cAMP combines with an enzyme that activates another enzyme which acts on the target substance (e.g., in the case of glucagon, this final enzyme converts glycogen into glucose). • Receptors are coupled to special proteins (G) that mediate intracellular events (all G proteins have 3 different subunits). The G protein receptor family includes glucagon, oxytocin, and vasopressin receptors. • When the hormone-receptor complex binds to G, G in turn activates adenylate cyclase, which in turn stimulates that formation of cyclic adenosine monophosphate, or cAMP. • When formed in response to a hormone-receptor bind, cAMP is referred to as the 2nd messenger (the hormone is the 1st messenger) • Once formed, cAMP can combine with an enzyme called protein kinase A (PKA), an enzyme that in turn activates (phosphorylates) another enzyme called phosphorylase kinase in a variety of cells. • For example, phosphorylase kinase A breaks down glycogen into glucose to provide intracellular energy. glycogen

29. Cellular and molecular mechanisms of hormone action--Steroid Hormones • Steroid hormones are fat soluble and move easily through cell membranes (as a result, these hormones are never stored but leave the cells in which they are produced almost immediately--nomadic). • The precursor to all vertebrate steroid hormones is cholesterol (made from acetate in the liver). • In the blood, steroid hormones must bind to water-soluble carrier proteins to increase their solubility and the ability of the blood to carry them to their target tissues. • Upon arrival, they dissociate from their carrier proteins and diffuse through the cell membrane into the cytoplasm of the target cell, where they bind to cytoplasmic receptors. • The steroid-receptor complex is transported into the cell nucleus, where it binds to DNA sequences called hormone response elements and then either stimulates or inhibits the transcription of specific mRNA. • The precise mechanism through which binding to the hormone response elements occurs is unknown. • The transcribed mRNA migrates to the cytoplasmic rough endoplasmic reticulum, where it is translated into specific proteins or enzymes that produce the physiological response (much more on this later). • Note: changes in the types of proteins a cell makes (aka, the gene products) can be observed within 30 minutes of hormone stimulation. cholesterol

30. The major vertebrate hormones • Protein & peptide hormones: • Make up the majority of hormones • Protein hormones that are only a few amino acids in length are called peptide hormones (larger ones called protein or polypeptide hormones) • Include: insulin, the glucagons, the neurohormones of the hypothalamus (monoamines), the hormones of the anterior pituitary, inhibin, calcitonin, parathyroid hormone, the GI hormones, leptin, and the posterior pituitary hormones • These hormones are blood-soluble (they don’t need a carrier protein to travel to their target cells, as do steroid hormones). However, they may bind with other blood plasma proteins • The metabolism of a hormone is reported in terms of its half-life, which is the amount of time required to remove half of the (radioactively tagged) hormone from the blood • Generally, larger protein hormones have longer half-lives (e.g., growth hormone with 200 amino acids has a half life of 20-30 mins, whereas thyroid releasing hormone has 3 amino acids and a half life of fewer than 5 minutes). Insulin (not Europe) Human growth hormone Too much human growth hormone (tumor on the anterior pituitary)

31. Hypothalamic hormones • The peptide hormones secreted by the hypothalamus are best thought of as a special class of neurotransmitters that act on a variety of cells in the anterior pituitary. • Five releasing hormones and one inhibiting hormone have been isolated. • TRH--thyrotropin-releasing hormone • GHRH--growth hormone-releasing hormone • GnRH--gonadotropin-releasing hormone • MSH--melanotropin-releasing hormone • CRH--corticotropin-releasing hormone • Somatostatin--growth hormone-inhibiting hormone • Peptide and protein hormones vary in amino acid sequence (and vary by species--e.g., GnRH possesses a different sequence in frogs than in horses; horse GnRH will not affect a frog’s reproductive function (although salmon calcitonin is used in humans to promote bone mineralization). Hormones differ in species specificity. Notice inter-species discrepancies in the 2nd position (glycine), 8th position (methionine),10th, 11th, and many others moving forward. Calcitonin is a thyroid hormone. It lowers blood levels of calcium by inhibiting calcium release from bone. Interestingly, it is regulated by blood calcium levels, not by pituitary hormones.

32. Anterior Pituitary hormones • All protein hormones, all, ranging in length from 39 to 220 amino acids. • Anterior pituitary is composed of three types of cells: • Acidophils (these cells stain readily with acidic stains) • Basophils (stain readily with basic stains) • Chromatophils (do not take up either acidic or basic stains) Basophil-secreted: • Lutenizing hormone (LH), follicule-stimulating hormone (FSH), and thyroid-stimulating hormone (TSH) are secreted by basophils. • LH and FSH and controlled by hypothalamic GnRH. TSH is controlled by hypothalamic TRH. • All consist of 200-220 amino acids and have molecular weights of 25-35K daltons. • Note: LH and FSH are known as gonadotropins because in response to GnRH, they stimulate the production of steroids in the gonads. Acidophil-secreted: • Growth hormone (GH) and prolactin (PRL) • TRH stimulates PRL secretion; hypothalamic dopamine inhibits. GHRH and Somatostatin control GH. • 190-220 amino acids in length. • GH shows fair amount of species specificity. • GH stimulates body growth INDIRECTLY (does not induce skeletal growth). • Stimulates production of growth-regulating substances--somatostedins--by the liver and kidneys. Somatostedins cause bone to take up sulfates leading to growth. • GH also stimulates protein synthesis, fat mobilization,and hyperglycemia (because of its anti-insulin properties). • Prolactin (PRL) best known for promoting lactation in female mammals, but in fact PRL has hundreds of physiological functions. Was originally called luteotropic hormone because its 1st known function was to promote corpus luteum function in rat ovary. • PRL functions can be broken down into 5 basic classes: • Reproduction • e.g., corpora lutea formation/maintenance • Growth and development • e.g., second metamorphosis of salamanders from terrestrial to aquatic adult • Water and electrolyte balance • e.g., certain minnows can migrate between seawater and fresh water. They are euryhaline. • Maintenance of integumentary system (external body covering) • e.g., pigeons and doves feed their young crop milk, which is secreted by the PRL-developed crop sac. • Actions on steroid-dependent target tissues or synergisms with steroids to affect target tissues. • e.g., PRL necessary to maintain LH receptors in the testes of mammals • Note: Elevated PRL levels appear to mediate paternal behavior in the California mouse and marmoset (reductions in testosterone have the same effects in the marmoset). Humans? Salt marsh killfish actual crop milk crop sac

33. Anterior Pituitary hormones (cont.) chromatophil secreted • ACTH (Adrenocorticotropic hormone) secreted by the chromatophils. • 39 amino acids; 4500 daltons in weight • ACTH released in response to CRH from the hypothalamus; stimulates the adrenal cortex to secrete mineralocorticoids and glucocorticoids (including cortisol, the primary glucocorticoid, which may feed back to control ACTH release. This conclusion is based on an increase in ACTH secretion after adrenalectomy.) Relationship between CRH, ACTH, and adrenals

34. Posterior Pituitary hormones • Two peptides, oxytocin and vasopressin, are released from the posterior lobe of the pituitary in mammals. • Oxytocin regulated by electrical activity of the oxytocin cells in the hypothalamus. Vasopressin secreted in response to increased osmotic pressure in the heart, veins, and carotid arteries. Oxytocin (Greek: “quick birth”) influences reproductive function • Important in birth: • Causes uterine contractions (synthetic oxytoxin--Pitocin--used to induce labor) • Causes the letdown reflex (in response to sensory stimulation of the nipples, oxytocin is released, travels to the mammary glands, which contract upon exposure, causing milk letdown--and because of prior associations--cry of a hungry baby, or the sound of a milking machine (in cows) produces the same mechanism. • Behaviorally: • oxytocin injected into the cerebrospinal fluid causes spontaneous erections in rats • In the Prairie Vole, oxytocin released into the brain of the female during sexual activity is important for forming a monogamous pair bond with her sexual partner • Sheep and rat females given oxytocin antagonists after giving birth do not exhibit typical maternal behavior. By contrast, virgin female sheep show maternal behavior towards foreign lambs upon cerebrospinal fluid infusion of oxytocin • Crossing the placenta, maternal oxytocin reaches the fetal brain and induces a switch in the action of neurotransmitter GABA from excitatory to inhibitory on fetal cortical neurons. This silences the fetal brain for the period of delivery and reduces its vulnerability to hypoxic damage Vasopressin (aka antidiuretic hormone--ADH--or arginine vasopressin--AVP--acts to retain water in four-footed vertebrates. • Rate of filtration in the kidneys slows in response to ADH, resulting in water retention. • ADH has hypertensive effects during serious blood loss--blood vessels constrict in response to severe hemorhage, slowing blood flow. • Behaviorally, vasopressin seems to induce the male to become aggressive towards other males.

35. Hypothalamic hormones GHRH GnRH MSH TRH CRH Somatostatin Pituitary Hormones Anterior GH (GHRH) FSH, LH (GnRH) TSH, PRL (TRH) ACTH (CRH) Posterior Oxytocin (hypothalamic oxytocin cells) Vasopressin (aka, ADH, AVP--increases in osmotic pressure). Review

36. Some other protein hormones

37. The Thyroid hormones • Thyroid gland releases its hormones in response to TSH stimulation from the anterior pituitary • There are two biologically active thyroid hormones, both derived from a molecule called thyroglobulin • T3, also known as tri-iodothyronine and T4, thyroxine have three general effects: • Increases metabolism--generally, thyroid activity is greater in the winter. • Growth and differentiation--closely related to actions of GH; in fact, effects probably represent permissive actions on GH target cells • permissive effects occur when one hormone induces receptor production for a second hormone. These effects are common. • Behavioral effects--insufficient production can affect CNS development, causing cretinism; insufficient production can also delay sexual maturation Endemic cretinism in the Democratic Republic of Congo. Four inhabitants aged 15-20 years : a normal male and three females with severe longstanding hypothyroidism with dwarfism, retarded sexual development, puffy features, dry skin and hair and severe mental retardation

38. The GI hormones • Three major gastrointestinal hormones: • Secretin • Released by the duodenum (1st segment of small intestine). Stimulates pancreas to produce water and bicarbonate, which aid in digestion. Also stimulates liver to produce bile. • Gastrin • Released by the stomach. Stimulates insulin release, smooth muscle contractions of gut, gallbladder, and uterus. • Cholecystokinin (CCK) • Released by duodenum. Causes pancreas to secrete digestive enzymes; causes gallbladder to contract and release bile

39. The Pancreatic hormones • Insulin • Only known hormone in the animal kingdom that can lower blood sugar (many hormones act to raise blood glucose levels). All cells (except for CNS cells) have insulin receptors. When an insulin receptor is activated, glucose is taken up into the cell, used, or stored as glycogen in muscle or fat cells. • Glucagon • Travels to the liver, where it breaks down stored glycogen (glycogenolysis), serving to increase blood levels of glucose. It acts in opposition to insulin.

40. Other peptide hormones • Enkephalins • Possibly involved in the stress response; released by the adrenal medulla • Inhibin • Blocks secretion of FSH and aromatase (an enzyme that converts estrogens from androgens); released by the gonads (testes and ovaries). See figure at right. • Activin • Directly stimulates aromatase activity in the gonads (opposite of inhibin?); released by the gonads • Relaxin • Softens pelvic ligaments during pregnancy to allow the large head of the fetus clear passage through the vaginal canal during birth; released by the ovaries Female mouse on top left is a transgenic mouse used to explore the potential effects of excess inhibin on the reproductive axis. The inhibin subunit protein was overexpressed in transgenic mice. The transgene is expressed in numerous tissues and levels of inhibin are highly elevated compared to control mice, leading to a decrease in serum FSH, and an increase in testosterone and serum LH. Activin levels are also somewhat depressed. The female mice are subfertile and have very small litters. This is a consequence of decreased ovulation, probably secondary to alterations in FSH and LH. Most interestingly, female mice that carry this transgene develop several unique ovarian pathologies, including distension of the bursal sac, the presence of large fluid-filled cysts, and the presence of atypical follicles that contain multiple oocytes (Figure 3).

41. The Steroid hormones • All steroids have a common chemical structure--three six-carbon rings plus one conjugated five carbon ring • The precursor to all steroid hormones is cholesterol (produce from acetate in the liver) • Recall that steroid hormones are fat-soluble and move easily through cell membranes (slide 25) • Further recall that in circulation, they must bind to water-soluble carrier proteins (slide 25) • Three major classes of steroid hormones: • Progestins/Corticoids • Androgens • Estrogens cholesterol

42. The C21 steroids: Progestin & Corticoids Two types of C21 hormones--progestins and corticoids • In response to anterior pituitary signals, cholesterol is converted into various steroid hormones in the adrenals • P450-linked side chain cleaving enzyme, or desmolase cuts cholesterol down intopregnenolone, which is a progestin and is the precursor to all other steroid hormones • Pregnenolone is a prohormone. • A prohormone is a substance that can act as a hormone and can be converted into another hormone with different endocrine properties • Progestins are named for their pregnancy-maintaining effects. Progesterone is important in maintaining pregnancy and in the initiation and cessation of mating behaviors (also, perhaps attachment) Two types of corticoids: glucocorticoids and mineralocorticoids • The glucocorticoids are involved in carbohydrate metabolism and are released under stress; the two primary are corticosterone and cortisol. All reptiles and birds, as well as rats and mice secrete corticosterone; the primary glucocorticoid in primates is cortisol • Aldosterone is the most important mineralocorticoid. Primarily responsible for retaining sodium and excreting potassium

43. The C19 steroids: The Androgens Progestins are precursors to all androgens. Enyzmes found in the gonads convert pregnenolone to several different androgens (from Greek andros for man). • Most biologically important are testosterone, androstendione, and dihydrotestosterone (DHT) • Produced in the Leydig cells of the testes; Sertoli cells are source of the androgen-binding proteins that carry androgens through the blood • Another androgen, DHEA (and DHEA-S) are produced in the adrenal cortex. • Relatively weak • Physio functions unknown • DHEA replacement may ameliorate certain aging effects (increased muscle mass, decreased fat mass) • Physiological functions of androgens • Spermatogenosis • Maintenance of the genital tract • Maintenance of the accessory sex organs (prostate et al.) • Secondary sex characteristics (body hair in humans, comb size in roosters, antler growth in deer) • Behavioral functions • Courtship • Copulation • Aggression • Dominance • Many other social behaviors (just beginning to be discovered) • Metabolic functions • Increase in muscle mass due to protein metabolism • Increase in respiration • At supraphysiologic levels, hypertrophy of organ systems due to widespread androgen receptor density (e.g., liver, heart, kidneys). Thus, chronic exposure to supraphysio levels can lead to a reduction in function • Reproductive function often compromised at supraphysio (along with testicular shrinkage due to downregulation of Leydig and Sertoli tissues)

44. The C18 steroids: The Estrogens Androgens are the precursors of all estrogens just as progestins are the precursors of all androgens Estrogen means “producing” (coined 1927) • Enzymes in the ovaries convert testosterone and androstendione to estrogen by a process called aromatization (because removal of the 19th carbon results in an aromatic compound). • Biologically significant estrogens are estradiol, estrone, and estriol. • Note that the ovaries produce androgens, which are then aromatized into estrogens. • Sometimes excess androgens are produced or insufficient enzymes are present and the female is masculinized. • Conversely, if high levels of enzymes that convert androgens to estrogens are present in the testes, then estrogens will be secreted into the blood and the male will be feminized. DHT cannot be aromatized (why is this important?) • Physiolgical functions • Initiate corpora lutea formation • Uterine mass density (estrogen levels positively correlated with . . .) • Development of secondary sex characteristics • Metabolic functions • Water retention • Calcium metabolism (bone mass increases in the presence of estrogen) • Behavioral functions • Very important in maternal aggression and sexual behavior (e.g., in combination with progesterone, prolactin, oxytocin, estradiol induces rats to behave maternally in the presence of pups). • Progesterone is primary in maternal aggression, but estrogens may play a role. Maternal aggression in women has not been studied. Many other social behaviors (just beginning to be discovered)

45. Androgens & Estrogens are not sex-linked • All males produce estrogens and progestins • All females produce androgens • The sex difference in circulating hormone levels is due to levels of gonadal enzymes. • Testes have more enzymes for making androgens and less aromatase than do ovaries • Ovaries produce high concentrations of androgens but these are easily aromatized into estrogens in the ovaries. These can be converted back into androgens, but this is an energetically expensive reaction • Recall that the adrenals produce sex hormones • Gene mutation(s) can lead to enzyme deficiencies which in turn can lead to an ovary or adrenal gland producing large quantities of androgens. See example at right Congenital andrenal hyperplasia (lacking the enzyme to metabolize cortisol and aldosterone, and as a result, produce too much androgen. This is a female with an extremely virilized clitoris.

46. Review FYI, no such medically recognized condition as adrenal fatigue.

47. The monoamines • Derived from a single amino acid • Two classes • Catecholamines • Indole amines Adrenal medulla monoamines • epinephrine • norepinephrine • dopamine • All derived from Tryosine • Released in response to sympathetic neural signals (evoked by stress, exercise, low temp, anxiety, emotionality, and hemorrhage). In humans, norepi and epi are released at a ratio of 1:4 • Actions: • Increase heart rate • Vasoconstriction of deep arteries and veins • Dilation of skeletal and liver blood vessels • Increased glycolysis • Increased blood glucagon concentrations and decreased insulin secretion Pineal gland indole amines • Serotonin (5-HT) • Melatonin • Derived from tryptophan • 5-HT levels high in the pineal during the day, but low at night as 5-HT is converted into melatonin • Actions: • The success of the SSRIs (e.g., prozac) work in depressives who lack the normal sensitivity to serotonin and thus benefit from the delay in reuptake that the SSRI provides. In essence, the serotonin has multiple chances to be recognized by the malfunctioning serotonin receptors due to lack of reuptake. • Low levels of 5-HT associated with aggression in many mammalian species • Melatonin involved in regulation of puberty onset and seasonal organization of breeding • Also, implicated in maintenance of biological rhythms (circadian, diurnal, seasonal) The brains of baby rhesus monkeys who endured high rates of maternal rejection and mild abuse in their first month of life produced less serotonin. Low levels of serotonin are linked to anxiety and depression and impulsive aggression in both humans and monkeys.

48. Regulation • Negative feedback • Example: • GnRH is released from hypothalamus • Gonadotropins are released from anterior pituitary • Steroid and gamete production in gonads is stimulated • Resulting steroid hormones turn off GnRH production in the hypothalamus, which shuts down gonadotropin release • Up-regulation • Increase in PRL stimulates production of more PRL receptors • Back to point raised on Slide #11 (many hormones have a long latency of action ). Up-regulation is the result of protein synthesis, and can take weeks for the necessary quantity of receptors to accommodate to the higher hormone levels (case in point--action of certain anti-depressives require a substantial increase in # of receptors to show effects; must wait on receptor synthesis). • This point also explains the failure of short-term hormone experiments (insufficient receptors to accommodate the sudden supra-physio hormone load). • Down-regulation • High insulin concentrations reduce the number of insulin receptors

49. So, WHY should social-personality psychologists study hormones? • Tiny handful of folks doing this stuff • Upside potential is unlimited • Grant $$ is untapped • Prostate cancer example • Opportunities for collaborations are great • Behavioral endocrinology is an interdisciplinary approach • fMRI? • Powerful tool to correlate cortical location, and it should be seen as one tool in the toolbox, just as self-reports should be seen as one tool among many. Other tools are critical if personality and social behavior are to be understood • So, what about the stuff that doesn’t or cannot get scanned--the limbic system, the amygdala, the hippocampus, the midbrain, the thalamus, the hypothalamus, the pituitary, and of course, the medulla oblongata