Unless otherwise noted, lectures are held the second Tuesday of each month at 4:00 PM in 341 Bardeen.

September 14th - The Regeneration GAP: Differences in gap43 Gene Regulation during Zebrafish CNS Development and Regeneration

Ava Udvadia
Department of Biological Sciences
University of Wisconsin-Milwaukee

Fish, unlike mammals, have the capacity for functional adult CNS regeneration, which is due, in part, to their ability to express axon growth-related genes in response to nerve injury. We have identified differences in gene regulatory elements between fish and mammals in one such axon growth gene, gap43, which is expressed in regenerating CNS neurons in adult fish, but not in adult mammals. Gap43 gene expression in neurons is associated with periods of axon growth and guidance during development of the nervous system and following nerve injury in neurons capable of regeneration. We previously demonstrated that the cis-regulatory elements of gap43 needed for developmental expression are not sufficient for expression during regeneration in the zebrafish. Here we have identified a 3.6 kb promoter fragment that can promote reporter gene expression in the nervous system during both development and regeneration. This fragment contains proximal promoter regions that are conserved across vertebrate species, and distal regions that are found in only in vertebrates capable of CNS regeneration (e.g. fish), but not in those that display limited CNS regeneration (e.g. birds and mammals). We demonstrate that the proximal promoter regions are sufficient for developmental expression, but not for regenerative expression, while the distal promoter regions are necessary for regenerative expression. These findings raise the possibility that the selective loss of key regulatory sequences during evolution may contribute to the differences in gap43 gene expression that occur among vertebrates following CNS injury.

October 12th - Nonlinear Optical Probes of Ovarian Cancer

Paul Campagnola
Departments of Biomedical Engineering and Medical Physics
University of Wisconsin-Madison

Remodeling of the extracellular matrix (ECM) has been implicated in ovarian cancer, and we hypothesize that these alterations may provide a highly sensitive optical marker of early disease. For this investigation we use Second Harmonic Generation (SHG) imaging microcopy to study changes in the structure of the ovarian ECM in human normal and malignant ex vivo biopsies. The normal and malignant tissues have highly different collagen fiber assemblies, where collectively, our findings show that the malignant ovaries are characterized by lower cell density, denser collagen, as well as higher regularity at both the fibril and fiber levels. This further suggests that the assembly in cancer may be comprised of newly synthesized collagen as opposed to modification of existing collagen. Because invasion and metastasis of human ovarian cancer are thought to be synchronous, it is crucial to understand the cancer cell adhesion, migration, and detachment dynamics. We use multiphoton excited photochemistry to fabricate crosslinked laminin nanofibers to investigate these processes as a function of metastatic potential. The migration rates increase with increasing metastatic potential, and the more invasive cells are less rigid and more weakly adhered to the nanofibers. The extent of directed migration also depends on the cell polarity and focal adhesion expression. For the invasive cells, these findings are similar to the integrin- independent ameboid-like migration seen for polar cells in collagen gels. Additionally, we find that all the cells display enhanced directed migration on crosslinked laminin gradients. Moreover, the migration speed and directionality depends not only on the local concentration and slope of the gradient but also on the cell polarity. Collectively, the results suggest that contact mediated migration, haptotaxis, as well as decreased adhesion may be operative in metastasis of ovarian cancer in vivo.

November 9th - Order and Disorder in the Emotional Brain
Note Time Change: 4:15PM

Richard Davidson
Department of Psychology
University of Wisconsin-Madison

Emotions are at the core of human personality, they define each person’s uniqueness and they shape resilience and vulnerability to adversity. Perhaps the single most salient characteristic of emotion is the variability across individuals in how each responds to emotional cues and challenges. This variability is termed “affective style.” Different parameters of affective style can be objectively measured and are instantiated in different underlying neural circuits. Activation patterns assessed with neuroimaging, particularly those involving interactions between sectors of the prefrontal cortex and subcortical structures implicated in emotion, are related to different parameters of affective style and are consistent over time within individuals. Specific patterns of brain activity are related to vulnerability to particular types of disorders. Moreover, patterns of central brain function are related to peripheral biological systems that play a role in physical health and illness. Despite their consistency over time within individuals, these patterns of neural activity are not immutable to change but rather can be transformed through systematic mental training such as meditation. The literature on neuroplasticity provides a framework for understanding these changes. This latter body of evidence supports the view that happiness, well-being and emotional balance are best regarded as the product of trainable skills.

December 14th - Molecular Regulation of Synapse Assembly and Number

Kate O'Connor-Giles
Department of Genetics
University of Wisconsin-Madison

The neural circuits that mediate nervous system function depend on the proper assembly and coordinated growth of synaptic connections between neurons and their targets. Our emerging understanding of a number of neurodevelopmental disorders suggests that aberrant synapse number, whether too few or too many, leads to cognitive impairments. Thus, the identification and characterization of molecules that control the extent of synaptic growth are critical to understanding normal neural function and our ability to treat a variety of neurological disorders. In Drosophila and mammals, a retrograde (postsynaptic to presynaptic) bone morphogenic protein (BMP) growth signal promotes synaptogenesis. We have found that the level of BMP signaling at the Drosophila neuromuscular junction (NMJ) regulates the extent of synaptic growth and identified an endocytic mechanism for modulating signal levels presynaptically. Through genetic screens at the Drosophila NMJ, we have identified and are functionally characterizing additional molecules and mechanisms that regulate BMP-dependent synaptic growth. We are taking a similar approach to understand the molecular regulation of synapse assembly. Effective neurotransmission depends on the precise and coordinated development of presynaptic active zones (AZs) that facilitate synchronous neurotransmitter release in direct apposition to postsynaptic densities (PSDs) comprising the neurotransmitter receptors and ion channels that mediate synaptic responses. Our findings illustrate the feasibility of utilizing the genetically tractable Drosophila NMJ as a model synapse to carry out functional analysis of synapse assembly in vivo.

January 18th - Seeing Cells in Action with Photoactivatable Fluorescent Proteins
Note Date (3rd Tuesday in January) and Location (140 Bardeen)

Jennifer Lippincott-Schwartz
Chief, Section on Organelle Biology

Photoactivatable fluorescent proteins (PA-FPs) are molecules that switch to a new fluorescent state in response to activation to generate a high level of contrast. Several types of PA-FPs have been developed, including PA-FPs that fluoresce green or red, or convert from green to red in response to activating light. The optical ‘‘highlighting’’ capability of PA-FPs has led to the rise of novel imaging techniques providing important new biological insights. These range from in cellulo pulse-chase labeling for tracking subpopulations of cells, organelles or proteins under physiological settings, to super-resolution imaging of single molecules for determining intracellular protein distributions at nanometer precision. The use of PA-FPs in super-resolution imaging of single molecules is a rapidly emerging field of microscopy that improves the spatial resolution of light microscopy by over an order of magnitude (10-20 nm resolution). It is based on the controlled activation and sampling of sparse subsets of photoconvertible fluorescent molecules whose illumination centroids are fitted and then summed into a final super resolution image, revealing the complex distribution of dense populations of molecules within subcellular structures with nanometer precision. The full potential of PA-FPs in conventional, diffraction-limited and super-resolution imaging is only beginning to be realized. Here, I discuss the diverse array of PA-FPs available to researchers and the new imaging techniques they make possible for unraveling long-standing biological questions.

February 8th - Single-Cell, Real-Time Observation of the Attack of the Human Antimicrobial Peptide LL-37 on Live E. coli
Note Location (5235 MSC)

James Weisshaar
Department of Chemistry
University of Wisconsin-Madison

LL-37 is a natural, cationic antimicrobial peptide present on human skin and lung tissue. It is thought to halt bacterial cell growth and eventually kill cells by degrading the barrier function of the cell membrane, but there is little direct mechanistic evidence. In the lungs of cystic fibrosis patients, LL-37 apparently loses its ability to kill bacteria. Two-color fluorescence microscopy enables us to correlate in time the onset of various bacterial “symptoms” with the halting of cell growth. In the K-12 strain of the Gram negative E. coli, growth halts when LL-37 translocates across the outer membrane to reach the thin periplasmic space, long before the cytoplasmic membrane is penetrated. The growth-halting mechanism may well be related to disruption of biosynthesis of new, curved cell wall during septation. Single-molecule counting experiments suggest that LL-37 penetrates the outer membrane at surface peptide/lipid ratios much smaller than those required to form pores in model synthetic lipid bilayers. Time permitting, we will also present preliminary results for the attack of LL-37 on the Gram positive B. subtilis.

March 8th - How the Brain Makes Up the Mind: The Biology of Personality and Decision-Making
Note Location (5235 MSC)

Michael Koenigs
Department of Psychiatry
University of Wisconsin-Madison

Why do people behave the way they do? Humans are clearly not 100% ‘rational’ creatures whose behaviors are strictly determined by universal social, moral, or economic laws. We each have our own particular set of beliefs, values, attitudes, fears, desires, and temperament—in short, we all have unique personalities. At some level, all of human behavior can be viewed as an output of the human nervous system, so is there a brain area that controls our individual personalities? If so, where is it and how does it work? This talk will describe studies on brain-damaged patients who have undergone dramatic changes in personality as a result of their brain injuries. These studies have identified a particular area of the brain—the ventromedial prefrontal cortex—that is especially critical for controlling emotion, decision-making, and social behavior. The knowledge gained from these lesion mapping studies may be applied to better understand illnesses such as depression, anxiety, and psychopathy.

April 12th - Linking Sensation to Action in the Brain of Behaving Drosophila
Note Location (5235 MSC)

Vivek Jayaraman
Janelia Farm Research Campus
Howard Hughes Medical Institute

We are interested in understanding basic principles of sensorimotor circuit function. We would like to understand how neural circuits integrate multisensory information with motor output to enable directed behavior. To this end, we have developed a novel preparation that allows stable physiology from identified neurons in the central brain of tethered (head-fixed) behaving Drosophila melanogaster. In our first study with the new setup, we performed two-photon calcium imaging using the genetically encoded calcium indicator, GCaMP3.0, expressed in the lobula plate tangential cells (LPTCs) in tethered walking (on an air-supported ball) and flying flies during optomotor behavior. When presented with horizontally moving visual patterns, flies displayed robust compensatory turning on the ball and in flight. Neurons of the horizontal system (HS)—a subgroup of LPTCs thought to be involved in optomotor behavior—responded with strong calcium transients correlated with the simultaneously recorded angular rotation of the fly. HS neurons, which had previously been recorded in fixed preparations, displayed stronger calcium transients in response to motion stimuli when flies were walking or flying rather than resting, and the strength of their responses was correlated with the fly’s own speed. Moreover, HS neurons showed a greater response gain for higher temporal frequency motion stimuli during movement, shifting their temporal frequency optimum towards higher speeds. When an animal moves through its environment, its retina is exposed to higher image speeds due to self-motion. Thus, state-dependent modulation of HS neuron tuning in the Drosophila visual system may constitute a mechanism to facilitate processing of faster retinal image shifts in behavioral contexts where these speeds of visual motion are relevant for course stabilization.

In addition to two-photon imaging experiments, I will also discuss tools we have developed to record electrical activity of identified populations of neurons in the fly brain using multi-electrodes and optogenetics. In ongoing experiments, we are now using two-photon imaging and electrophysiology to explore the functioning of deeper brain regions in the fly brain in the context of sensorimotor orienting behavior.