How the Vertebrate Brain Regulates Behavior

by Donald Pfaff

  ______. 1995. “Functional Analyses of α1-Adrenoceptor Subtypes in Rat Hypothalamic Ventromedial Nucleus Neurons.” European Journal of Pharmacology 282: 199–206.

  ______. 1998. “Mapping of Neural and Signal Transduction Pathways for Lordosis in the Search for Estrogen Actions on the Central Nervous System.” Behavioral Brain Research 92: 169–180.

  Kow, L.-M., Y. F. Tsai, N. G. Weiland, B. S. McEwen, and D. W. Pfaff. 1995. “In Vitro Electro-Pharmacological and Autoradiographic Analyses of Muscarinic Receptor Subtypes in Rat Hypothalamic Ventromedial Nucleus: Implications for Cholinergic Regulation of Lordosis.” Brain Research 694: 29–39.

  Kow, L.-M., G. D. Weesner, and D. W. Pfaff. 1992. “Alpha 1-Adrenergic Agonists Act on the Ventromedial Hypothalamus to Cause Neuronal Excitation and Lordosis Facilitation: Electrophysiological and Behavioral Evidence.” Brain Research 588: 237–245.

  Krebs, C., E. Jarvis, J. Chan, J. Lydon, S. Ogawa, and D. W. Pfaff. 2000. “A Membrane-Associated Progesterone Binding Protein, 25-Dx, Is Regulated by Progesterone in Brain Regions Involved in Female Reproductive Behaviors.” Proceedings of the National Academy of Sciences of the United States of America 97: 12816–12821.

  Lauber, A. H., C. V. Mobbs, M. Muramatsu, and D. W. Pfaff. 1991. “Estrogen Receptor mRNA Expression in Rat Hypothalamus as a Function of Genetic Sex and Estrogen Dose.” Endocrinology 129 (6): 3180–3186.

  Lauber, A. H., G. J. Romano, C. V. Mobbs, R. D. Howells, and D. W. Pfaff. 1990a. “Estradiol Induction of Proenkephalin Messenger RNA in Hypothalamus: Dose-Response and Relation to Reproductive Behavior in the Female Rat.” Brain Research: Molecular Brain Research 8: 47–54.

  ______. 1990b. “Estradiol Regulation of Estrogen Receptor Messenger Ribonucleic Acid in Rat Mediobasal Hypothalamus: An in Situ Hybridization Study.” Journal of Neuroendocrinology 2 (5): 605–611.

  Lauber, A. H., G. J. Romano, and D. W. Pfaff. 1991. “Sex Difference in Estradiol Regulation of Progestin Receptor mRNA in Rat Mediobasal Hypothalamus as Demonstrated by in Situ Hybridization.” Neuroendocrinology 53: 608–613.

  Lee, A. W., A. Kyrozis, V. Chevaleyre, L.-M. Kow, N. Devidze, Q. Zhang, A. M. Etgen, and D. W. Pfaff. 2008. “Estradiol Modulation of Phenylephrine-Induced Excitatory Responses in Ventromedial Hypothalamic Neurons of Female Rats.” Proceedings of the National Academy of Sciences of the United States of America 105 (20): 7333–7338.

  Lustig, R. H., D. W. Pfaff, and C. V. Mobbs. 1989. “Two-Dimensional Gel Autoradiographic Analysis of the Acute Effects of Estradiol on Protein Synthesis in the Female Rat Ventromedial Nucleus in Vivo.” Endocrinology 124: 1863–1869.

  Magariños, A. M., and D. W. Pfaff. 2016. “Sexual Motivation in the Female and Its Opposition by Stress.” In Behavioral Neuroscience of Motivation. Vol. 27 of Current Topics in Behavioral Neuroscience. Edited by E. H. Simpson and P. D. Balsam. Berlin: Springer, 35–49.

  Matsuda, K. I., H. Mori, B. M. Nugent, D. W. Pfaff, M. M. McCarthy, and M. Kawata. 2011. “Histone Deacetylation during Brain Development Is Essential for Permanent Masculinization of Sexual Behavior.” Endocrinology 152: 2760–2767.

  McCabe, J. T., J. I. Morrell, R. Ivell, H. Schmale, D. Richter, and D. W. Pfaff. 1986. “In Situ Hybridization Technique to Localize rRNA and mRNA in Mammalian Neurons.” Journal of Histochemistry and Cytochemistry 34: 45–50.

  McCarthy, M. M., S. P. Kleopoulos, C. V. Mobbs, and D. W. Pfaff. 1994. “Infusion of Antisense Oligodeoxynucleotides to the Oxytocin Receptor in the Ventromedial Hypothalamus Reduces Estrogen-Induced Sexual Receptivity and Oxytocin Receptor Binding in the Female Rat.” Neuroendocrinology 59: 432–440.

  Meisel, R. L., G. Dohanich, B. S. McEwen, and D. W. Pfaff. 1987. “Antagonism of Sexual Behavior in Female Rats by Ventromedial Hypothalamic Implants of Antiestrogen.” Neuroendocrinology 45: 201–207.

  Meisel, R. L., and D. W. Pfaff. 1984. “RNA and Protein Synthesis Inhibitors: Effects on Sexual Behavior in Female Rats.” Brain Research Bulletin 12: 187–193.

  ______. 1985. “Specificity and Neural Sites of Action of Anisomycin in the Reduction or Facilitation of Female Sexual Behavior in Rats.” Hormones and Behavior 19: 237–251.

  Mobbs, C. V., G. Fink, M. Johnson, W. Welch, and D. W. Pfaff. 1989. “Similarity of an Estrogen-Induced Protein and a Luteinizing Hormone Releasing Hormone Induced Protein.” Molecular and Cellular Endocrinology 62: 297–306.

  Mobbs, C. V., G. Fink, and D. W. Pfaff. 1990. “HIP-70: A Protein Induced by Estrogen in the Brain and LH-RH in the Pituitary.” Science 247: 1477–1479.

  Mobbs, C. V., R. E. Harlan, M. R. Burrous, and D. W. Pfaff. 1988. “An Estradiol-Induced Protein Synthesized in the Ventral Medial Hypothalamus and Transported to the Midbrain Central Gray.” Journal of Neuroscience 8: 113–118.

  Morgan, M., T. L. Dellovade, and D. W. Pfaff. 2000. “Effect of Thyroid Hormone Administration on Estrogen-Induced Sex Behavior in Female Mice.” Hormones and Behavior 37: 15–22.

  Nicot, A., S. Ogawa, Y. Berman, K. D. Carr, and D. W. Pfaff. 1997. “Effects of an Intrahypothalamic Injection of Antisense Oligonucleotides for Preproenkephalin mRNA in Female Rats: Evidence for Opioid Involvement in Lordosis Behavior.” Brain Research 777: 60–68.

  Ogawa, S., U. E. Olazabal, I. S. Parhar, and D. W. Pfaff. 1994. “Effects of Intrahypothalamic Administration of Antisense DNA for Progesterone Receptor mRNA on Reproductive Behavior and Progesterone Receptor Immunoreactivity in Female Rat.” Journal of Neuroscience 14: 1766–1774.

  Parsons, B., N. J. MacLusky, L. Krey, D. W. Pfaff, and B. S. McEwen. 1980. “The Temporal Relationship between Estrogen-Inducible Progestin Receptors in the Female Rat Brain and the Time Course of Estrogen Activation or Mating Behavior.” Endocrinology 107: 774–779.

  Parsons, B., N. J. MacLusky, M. S. Krieger, B. S. McEwen, and D. W. Pfaff. 1979. “The Effects of Long-Term Estrogen Exposure on the Induction of Sexual Behavior and Measurements of Brain Estrogen and Progestin Receptors in the Female Rat.” Hormones and Behavior 13: 301–313.

  Parsons, B., B. S. McEwen, and D. W. Pfaff. 1982. “A Discontinuous Schedule of Estradiol Treatment Is Sufficient to Activate Progesterone-Facilitated Feminine Sexual Behavior and to Increase Cytosol Receptors for Progestins in the Hypothalamus of the Rat.” Endocrinology 110: 613–619.

  Parsons, B., T. C. Rainbow, D. W. Pfaff, and B. S. McEwen. 1981. “Oestradiol, Sexual Receptivity and Cytosol Progestin Receptors in Rat Hypothalamus.” Nature 292: 58–59.

  ______. 1982. “Hypothalamic Protein Synthesis Essential for the Activation of Lordosis Behavior in the Female Rat.” Endocrinology 110: 620–624.

  Pfaff, D. W. 1973. “Luteinizing Hormone Releasing Factor (LRF) Potentiates Lordosis Behavior in Hypophysectomized Ovariectomized Female Rats.” Science 182: 1148–1149.

  ______. 1999. Drive: Neurobiological and Molecular Mechanisms of Sexual Motivation. Cambridge, MA: MIT Press.

  ______. 2005. Brain Arousal and Information Theory: Neural and Genetic Mechanisms. Cambridge, MA: Harvard University Press.

  Pfaff, D. W., L. Rosello, and P. Blackburn. 1984. “Proteins Synthesized in Medial Hypothalamus and Transported to Midbrain in Estrogen-Treated Female Rats.” Experimental Brain Research 57: 204–208.

  Pfaus, J. G., and D. W. Pfaff. 1992. “Mu-, Delta-, and Kappa-Opioid Receptor Agonists Selectively Modulate Sexual Behaviors in the Female Rat: Differential Dependence on Progesterone.” Hormones and Behavior 26: 457–473.

  Quiñones-Jenab, V., S. Jenab, S. Ogawa, R. A. Adan, J. P. Burbach, and D. W. Pfaff. 1997. “Effects of Estrogen on Oxytocin Receptor Messenger Ribonucleic Acid Expression in the Uterus, Pituitary, and Forebrain of the Female Rat.” Neuroendocrinology 65: 9–17.

  Quiñones-Jenab, V., S. Jenab, S. Ogawa, T. Funabashi, G. D. Weesner, and D. W. Pfaff. 1996. “Estrogen Regulation of Gonadotropin-Releasing Hormone Receptor Messenger RNA in Female Rat Pituitary Tissue.” Brain Research: Molecular Brain Research 38: 243–250.

  Quiñones-Jenab, V., S. Ogawa, S. Jenab, and D. W. Pfaff. 1996. “Estr
ogen Regulation of Preproenkephalin Messenger RNA in the Forebrain of Female Mice.” Journal of Chemical Neuroanatomy 12: 29–36.

  Rachman, I. M., D. W. Pfaff, and R. S. Cohen. 1996. “NADPH Diaphorase Activity and Nitric Oxide Synthase Immunoreactivity in Lordosis-Relevant Neurons of the Ventromedial Hypothalamus.” Neuroendocrinology 64: 357–363.

  Rachman, I. M., J. R. Unnerstall, D. W. Pfaff, and R. S. Cohen. 1998. “Regulation of Neuronal Nitric Oxide Synthase mRNA in Lordosis-Relevant Neurons of the Ventromedial Hypothalamus following Short-Term Estrogen Treatment.” Brain Research: Molecular Brain Research 59: 105–108.

  Romano, G. J., A. Krust, and D. W. Pfaff. 1989. “Expression and Estrogen Regulation of Progesterone Receptor mRNA in Neurons of the Mediobasal Hypothalamus: An in Situ Hybridization Study.” Molecular Endocrinology 3: 1295–1300.

  Romano, G. J., C. V. Mobbs, R. D. Howells, and D. W. Pfaff. 1989. “Estrogen Regulation of Proenkephalin Gene Expression in the Ventromedial Hypothalamus of the Rat: Temporal Qualities and Synergism with Progesterone.” Brain Research: Molecular Brain Research 5: 51–58.

  Romano, G. J., C. V. Mobbs, A. Lauber, R. D. Howells, and D. W. Pfaff. 1990. “Differential Regulation of Proenkephalin Gene Expression by Estrogen in the Ventromedial Hypothalamus of Male and Female Rats: Implications for the Molecular Basis of a Sexually Differentiated Behavior.” Brain Research 536: 63–68.

  Rothfeld, J. M., J. F. Hejtmancik, P. M. Conn, and D. W. Pfaff. 1987. “LHRH Messenger RNA in Neurons in the Intact and Castrate Male Rat Forebrain Studied by in Situ Hybridization.” Experimental Brain Research 67: 113–118.

  ______. 1989. “In Situ Hybridization for LHRH mRNA following Estrogen Treatment.” Brain Research: Molecular Brain Research 6: 121–125.

  Schumacher, M., H. Coirini, D. W. Pfaff, and B. S. McEwen. 1990. “Behavioral Effects of Progesterone Associated with Rapid Modulation of Oxytocin Receptors.” Science 250: 691–694.

  Scott, R. E., X. S. Wu-Peng, M. G. Kaplitt, and D. W. Pfaff. 2003. “Gene Transfer and in Vivo Promoter Analysis of the Rat Progesterone Receptor Using a Herpes Simplex Virus Viral Vector.” Brain Research: Molecular Brain Research 114: 91–100.

  Scott, R. E. M., S. Wu-Peng, and D. W. Pfaff. 2002. “Regulation and Expression of Progesterone Receptor mRNA Isoforms A and B in the Male and Female Rat Hypothalamus and Pituitary following Oestrogen Treatment.” Journal of Neuroendocrinology 14: 175–183.

  Scott, R. E. M., S. Wu-Peng, P. M. Yen, W. W. Chin, and D. W. Pfaff. 1997. “Interactions of Estrogen- and Thyroid Hormone Receptors on a Progesterone Receptor Estrogen Response Element (ERE) Sequence: A Comparison with the Vitellogenin A2 Consensus ERE.” Molecular Endocrinology 11: 1581–1592.

  Shivers, B., R. Harlan, J. Morrell, and D. W. Pfaff. 1983. “Absence of Oestradiol Concentration in Cell Nuclei of LHRH-Immunoreactive Neurones.” Nature 304: 345–347.

  Vasudevan, N., G. Davidkova, Y. S. Zhu, N. Koibuchi, W. W. Chin, and D. W. Pfaff. 2001. “Differential Interactions of Estrogen Receptor and Thyroid Hormone Receptor Isoforms on the Rat Oxytocin Receptor Promoter Leads to Differences in Transcriptional Regulation.” Neuroendocrinology 74: 309–324.

  Vasudevan, N., N. Koibuchi, W. W. Chin, and D. W. Pfaff. 2001. “Differential Crosstalk between Estrogen Receptors (ER-α) and (ER-β) and Thyroid Hormone Receptor Isoforms Results in Flexible Regulation of the Consensus ERE.” Brain Research: Molecular Brain Research 95: 9–17.

  Vasudevan, N., S. Ogawa, and D. W. Pfaff. 2002. “Estrogen and Thyroid Hormone Receptor Interactions: Physiological Flexibility by Molecular Specificity.” Physiological Reviews 82: 923–944.

  Vasudevan, N., Y. S. Zhu, S. Daniel, N. Koibuchi, W. W. Chin, and D. W. Pfaff. 2001. “Crosstalk between Oestrogen Receptors and Thyroid Hormone Receptor Isoforms Results in Differential Regulation of the Preproenkephalin Gene.” Journal of Neuroendocrinology 13: 779–790.

  Zhao, X., H. Lorenc, H., H. Stephenson, Y. J. Wang, D. Witherspoon, B. Katzenellenbogen, D. W. Pfaff, and N. Vasudevan. 2005. “Thyroid Hormone Can Regulate Estrogen-Mediated Transcription from a Consensus Estrogen Response Element In Neuroblastoma Cells.” Proceedings of the National Academy of Sciences of the United States of America 102 (13): 4890–4895.

  Zhou, J., A. W. Lee, Q. Zhang, L.-M. Kow, and D. W. Pfaff. 2007. “Histamine-Induced Excitatory Responses in Mouse Ventromedial Hypothalamic Neurons: Ionic Mechanisms and Estrogenic Regulation.” Journal of Neurophysiology 98 (6): 3143–3152.

  Zhu, Y. S., L. Q. Cai, X. You, Y. Duan, J. Imperato-McGinley, W. W. Chin, and D. W. Pfaff. 2001. “Molecular Analysis of Estrogen Induction of Preproenkephalin Gene Expression and Its Modulation by Thyroid Hormones.” Brain Research: Molecular Brain Research 91: 23–33.

  4

  GENES REGULATING BEHAVIOR

  Problem: My field was faced with questions about the reliability of studies using gene manipulation. Could we come up with a novel, reliable set of findings for a complete, normal mammalian behavior? How do genes regulate the reproductive behavior that we analyzed at the neuroanatomical, neurophysiological, and transcriptional levels in Chapters 1, 2, and 3? From genetics to physiology to behavior.

  Up to this point in our effort to show how a mammalian behavior is produced, I had spelled out the neuroanatomy of estrogen-binding neurons and the neurophysiology of the neural circuit that produces lordosis behavior. Because these estrogen-binding neurons serve estrogen-regulated transcription (Chapter 3), I now could use that family of facts to explore directly the impact of specific genes on reproductive behavior. That is, my field developed into applied genomics: a crescendo of neurobiological accomplishment with respect to reproductive (social) behavior as it subsumed genetic and epigenetic techniques. Thus, here I will relate how I used the chance to add up-to-date genetic and genomic technologies further to demonstrate exactly how estrogen-responsive neurons work in the lordosis neural circuitry to manage the behavioral side of reproduction. The findings provided the first and clearest gene / behavior causal relations available. I show how genetic alterations regulate reproductive behavior.

  Putting it another way, showing the estrogen-sensitive neurons that regulate our neural circuit for lordosis behavior and now showing how estrogen-sensitive genes regulate that behavior, we demonstrated for the first time how specific signaling chemicals from the body (hormones) acting on specific nerve cell groups regulate an entire instinctive mammalian behavior—a social behavior at that.

  Initial Uncertainties

  Keeping in mind Sir Peter Medawar’s phrase about “science being the art of the soluble,” I had focused our studies on a relatively simple, biologically crucial mammalian social behavior, calculating that it would provide a reasonable opportunity for a definitive explanation, an explanation with the kind of certainty approaching that of the physical sciences. That is, the intellectual paths that some scientists, including myself, have followed toward behavioral experimentation have traversed into physics as well as biology and other academic territories. Those who have studied physics may well be disposed to strive for demonstrations of universal laws, expressed quantitatively, in the proof of reliable stimulus–response connections. This endeavor can be hard because of the complexity of some stimuli and certain behavioral responses and because of the need to recognize and control the relevant environmental variables.

  The delineation of causal relations between particular genes and specific behaviors (Pfaff 2001) is even harder than purely behavioral research. The pleiotropy of individual genes and overlapping functions between genes makes for trouble—difficulties of analysis—to begin with. Our lack of understanding of the mechanisms for penetrance of dominant alleles makes it difficult to construct meaningful gene dose–response relationships. Interpretations of gene knockout data can stumble over unexpected compensations, effects on other gene products, and altered endocrine and neuronal feedback loops, as well as a lack of control over the temporal and spatial impact of the genetic manipulation.

  Nevertheless, as I will s
how, a significant body of mouse behavior genetics work that depends on modern molecular manipulations has emerged. When the experimenter chooses well-controlled stimuli and biologically important responses of enough simplicity, and their connections are driven by identified neuroendocrine or neurochemical influences of sufficient power, he or she can gain reliable, important knowledge. Therefore, it was surprising when John Crabbe at Oregon Health & Science University reported that he and his colleagues got somewhat different results on different campuses during analyses of mouse strain differences in behavior. More disturbing were the echoes in secondary sources that seemed to reinforce the old stereotype that many behavioral results are unreliable.

  Brutally clear are the effects of genes whose alterations lead to developmental pathologies in the cerebellum causing simple sensory-motor control abnormalities. A deletion within the RAR-related orphan receptor-α gene produces the staggerer mouse, showing abnormalities in cerebellar Purkinje cells associated with severe motor ataxia. This phenotype can be contrasted with the whole-body action tremors characteristic of the vibrator mouse. Here the pathology is due to a retroposon insertion in an intron of the phosphatidylinositol transfer protein α gene, preventing a normal accumulation of its RNA. Mutation of the subunit of a voltage-gated calcium channel could produce the tottering phenotype, which includes both ataxia and paroxysmal dyskinesis.

  So those motor abnormalities were clearly replicable, and that literature gave me the confidence to go forward to explain entire, biologically crucial, instinctive behaviors. I linked genes and consequent neurochemistry to physiology to behavior, and we can see exactly how gene / behavior causal relations can work out.

  The Gene for Estrogen Receptor α