Laboratory of Neurobiology and Behavior
Donald W. Pfaff
Laboratory of Neurobiology and Behavior
Donald Pfaff's new book was published in hardcover in August 1999 and paperback in September 1999. It is available at Amazon.com
Some of the lab's work can be summarized in four steps. First we worked on the localization of hormone target neurons in the brain and discovered estrogen-binding neurons in a limbic/hypothalamic system. The discovery initially was made in rat brain, but our work on fish CNS through monkey CNS showed it to be a general vertebrate system . We followed up the histochemical findings to demonstrate consequences of hormone binding for electrophysiological activity and neuronal growth. Secondly, we then worked out the first neural circuit for a vertebrate behavior, the estrogen-dependent lordosis behavior. The lordosis behavior circuit proved that it is possible to explain how mechanisms for a vertebrate behavior work. Third, we found hormone-dependent genes in the brain . Their induction by estrogenic hormones has temporal, spatial and gender specificities appropriate to reproductive behavior. Fourth, in turn, the products of some of these hormone-dependent genes are required for hormone-dependent lordosis behavior.
Taken together, these four findings showed that specific neurochemical reactions in specific parts of the brain determine a specific mammalian behavior .
Some of the Lab's New Work
- Transcriptional studies of how fast, membrane-initiated hormone effects facilitate later genomic effects. Parallel biophysical and behavioral studies.
- Microarray studies of steroid hormone effects on gene expression in specific brain regions of the adult; and of sex differences in the developing brain. Follow-up at RNA and protein levels. Predictions for functional genomic studies.
- Molecular, biophysical and behavioral studies of 'generalized arousal' transmitter actions on hypothalamic nerve cells. These include opioid peptides, histamine and norepinephrine.
- Exploring the mathematical structure of arousal components in mouse behavior. We are experimentally 'setting up' two forces on arousal, circadian and hunger, to study their interactions.
- Genetic and hormonal influences on brain arousal -- both sexual and generalized arousal in mice. We are breeding for 'high arousal' and 'low arousal' mouse lines. We have mouse models of diffuse brain damage that reduce arousal. Also we have a locus coeruleus-derived cell line for molecular analyses.
- Hormonal and genetic influences on the activation of neurons in the brainstem arousal system.
- Use of viral vectors to 'give back' missing genes in specific regions of the mouse brain, and to deliver siRNA to particular CNS regions.
Molecular Mechanisms for Behavior
We use molecular techniques to analyze: 1) how the mammalian brain manages specific natural behaviors; and 2) hormonal and genetic influences on generalized brain arousal. Some of this work can be done in nerve cell lines, but it is really necessary to study nerve cells in their normal synaptic context to see how, in the governance of behavior, the brain's special connectivity uses the types of molecular mechanisms seen in other tissues.
Advantageous for molecular studies, hormone effects on nerve cells build upon some of the best examples of eukaryotic transcription control. Steroid sex hormones and stress hormones have massive developmental effects, and in the adult brain they control a variety of natural behaviors. During development, for example, sex hormone actions around the time of birth determine behavioral sex differences, and early stress hormone exposure influences later responses to stress.
We have shown significant effects of estrogens on the transcription of the genes for the progesterone receptor and for the opioid peptide enkephalin. These hormone effects occur in specific parts of the brain and are required for normal reproductive behavior. They are strong in females but not males, again correlated with behavioral results. These transcriptional effects are due to the binding of estrogen receptors to their cognate DNA sequences, "estrogen response elements," in the promoters of the progesterone receptor and enkephalin genes. Other transcriptional systems that are hormone sensitive in the brain include the genes for oxytocin, the oxytocin receptor, the delta opioid receptor, the alpha adrenergic receptor, GnRH (LHRH) and the GnRH receptor. Microarrays have added many more candidate genes.
Interestingly, other transcription factors can interact with the estrogen receptor in the brain to influence estrogenic effects on gene expression and behavior. That is, thyroid hormone receptors, themselves transcriptionally active, can interfere with estrogen-dependent transcription and behavior. In doing so, thyroid hormones and their receptors bring reproductive controls into concordance with environmental signals, particularly environmental temperature.
In addition to explaining specific hormone-influenced behavior, we are uncovering mechanisms for fundamental brain arousal. We measure this in mice and influence it by genetic alterations.
The explosion in the number of interesting genetically altered mice is giving us new insights into the relationship between mammalian gene expression and behavior. Using estrogen receptor knockout mice, we showed that the effect of a specific gene on a specific behavior can depend upon the gender in which that gene is expressed as well as upon exactly when and where it is expressed. Functional genomics applied to these problems employ biophysical and behavioral assays in knockout mice. For us, that includes patch clamping of identified cells in mouse forebrain, with their molecular signatures determined by RT/PCR. In turn, the combination of mouse gene knockouts with neuropharmacology and antisense DNA technology offers the chance to open a new era, understanding mechanisms connecting gene expression to mammalian behavior.