School of Medicine Microscopy Seminar Series

UCSD - CNCB Large Conference Room

Refreshments will be served

Sept 10th 4PM - Nick Holland PhD

Scripps Institute of Oceanography, UCSD

A Serial Block-Face SEM Study of Nephrogenesis and Mouth Formation in Developing Amphioxus (a proxy for the basal chordate ancestor of the vertebrates)

Cephalochordates (commonly called amphioxus or lancelets), the most basal subphylum of Chordates, have been much studied for insights they  give into the origin of vertebrate structures. For over a century, the embryonic origin and the structure of the kidney of amphioxus has been controversial. Some biologists considered it the forerunner of the vertebrate pronephros, but other biologists likened it to the protonephridia of lower invertebrates. The nature of the amphioxus mouth is similarly contentious—some consider it homologous to the mouths of animals generally, some think it is really a modified gill slit, and some think it is a modified coelomoduct. Both kidney and mouth originate close together near the anterior end of an embryo in a tissue volume roughly 100 micrometers on a side. The action takes place on a scale that is too small to describe with light microscopy, but too large to study effectively with transmission electron microscopy. I used serial block-face scanning electron microscopy (SBSEM) to reconstruct in 3-D the important parts of amphioxus embryos at several developmental stages. Novel features of the work were the very large tissue volume studied and the low magnifications used. The results favor the pronephric nature of the amphioxus kidney and do not support the coelomoduct origin of the vertebrate mouth (Holland ND 2018, EvoDevo 9: article 16).

Imaging Technique Used:

Serial Block-Face Electron Microscopy

Sept. 11th 12PM - FLIM and STED Informational Seminar

Carlos Alonso - Leica Application Specialist

Sept 20th 4PM - Stephan Lange PhD

Associate Professor of Medicine, Division of Cardiovascular Medicine – UCSD

From Obscurity Into the Limelight: How Serial Block-Face Imaging Helped to Characterize the Heart-Failure Phenotype in a New Mouse Model of HFpEF

Muscle proteins of the obscurin protein family were shown to play roles for sarcomere organization, sarcoplasmic reticulum (SR) and sarcolemma architecture and function. However, their precise biological roles and involvement in cardiac diseases remain to be fully understood.

We set out to investigate the role that obscurin and its close homologue obscurin-like 1 (Obsl1) play for cardiac development and function. We generated and analyzed cardiac functions of obscurin, Obsl1 single, as well as obscurin/Obsl1 double-knockout mice using traditional and doppler trans-thoracic echocardiography, hemodynamics and P/V-loop studies. Changes to SR structure, protein content and ultrastructure were investigated by immunofluorescence, immunoblot, unbiased proteome and serial block-face electron microscopy analyses. SR-dependent changes to cellular calcium cycling were studied by imaging isolated cardiomyocytes.

Mice lacking obscurin and Obsl1 develop normally, but show dramatic changes to SR-architecture and function on the microscopic and proteome level. While obscurin has been shown to be important for SR-structure, our data reveal for the first time that Obsl1 has similar functions. Alterations to SR-structure are also reflected in dramatically reduced SR-volume and calcium cycling. Isolated double-knockout cardiomyocytes displayed reduced calcium amplitude (calcium release) and prolonged calcium re-uptake (tau-values). While systolic functions are near normal on the physiological level in controls, loss of obscurin/Obsl1 results in a profound relaxation defect in double knockout mice only, reflected by increased Tau and Min dP/dT-values from our hemodynamics analyses or elevated E/E' ratio. Intriguingly, the diastolic dysfunction is not accompanied by hypertrophy, increased fibrosis or changes to classical markers for cardiomyopathy (e.g. ANF), but changes on the metabolic level. Taken together, our data indicate that obscurin and Obsl1 are crucial for SR-structure, calcium storage and re-uptake. We propose that obscurin/Obsl1 double-knockout mice may serve as a new model to investigate age-dependent diastolic dysfunction and HFpEF.

Imaging Techniques Used:

Confocal Microscopy

Serial Block-Face Electron Microscopy

Oct. 2nd 12PM  Leong Chew PhD

Director of Janelila Farms Advanced Imaging Center

Imaging Life with the Emerging Frontiers in Spatial and Temporal Resolution at the Advanced Imaging Center, HHMI Janelia Research Campus

Visualizing and understanding complex biological processes demand the integrated efforts of biologists and physicists. As instrument developers continue to push the frontiers of what biologists can visualize with the latest technologies, the mission of the Advanced Imaging Center (AIC) is to make cutting edge imaging technologies developed at Janelia widely accessible, and at no cost, to scientists outside of Janelia, before the instruments are commercially available. Operating strategically at the interface of engineering and biological applications, the AIC is positioned to drastically reduce the time between instrument development and widespread use in the increasingly technology-intensive field of biology, thus positioned to empower investigators with tools currently not widely available elsewhere. In alignment with Janelia's philosophy of encouraging bold and risky science, the AIC welcomes proposals with high-risk-high-gain projects that may challenge the current paradigm. In fact, it serves as an ideal platform for researchers to test out their novel ideas with the emerging microscopy technologies, fully supported by Janelia's in-house imaging experts and research infrastructure. This talk will describe the cutting-edge science that has been made possible by this fleet of newly developed instruments, and the peer-review program through which non-Janelia scientists could access these technologies.  

Oct. 21st 4PM Chao-Wei Hung

Postdoctoral Fellow - Dr. Alan Saltiel Lab at UCSD

The Exocyst as a Gatekeeper for Insulin induces Glut4 Translocation

Glucose transport is the rate-limiting step by which insulin controls energy storage, and is mediated by the transporter Glut4. Despite years of study, the cellular machinery that controls insulin stimulated Glut4 translocation remains incompletely understood. Recent studies and preliminary data indicated that Glut4 translocation is initiated by a small G protein RalA, and the final steps are regulated by the phosphorylation of the exocyst complex by protein kinase TBK1. Our work examine the interaction of RalA and the exocyst components with respect to TBK1 activity in live cells by high-resolution time lapse microscopy. We aim to elucidate the temporal and spatial regulation of RalA and the exocyst during insulin signaling. Results from this study will delineate the sequential molecular events by which insulin controls the rate-limiting steps involved in Glut4 translocation to the plasma membrane in adipocytes, and will provide new insights into developing new treatment for type II diabetes.

Imaging Techniques Used:

Fluorescence Lifetime Imaging

TIRF Microscopy

Nov. 4th 4PM Matthew Shtrahman

Assistant Professor, UCSD Department of Neurosciences 

Microscopy Basics


All publications generated utilizing our resources must cite our grant: NS047101