ABSTRACT

First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping early-career researchers promote themselves alongside their papers. Jana Kroll and Özge Demet Özçete are co-first authors on ‘AP180 promotes release site clearance and clathrin-dependent vesicle reformation in mouse cochlear inner hair cells’, published in JCS. Jana is a Postdoc in the lab of Prof. Dr Christian Rosenmund at Institute of Neurophysiology, Charité Universitätsmedizin, Berlin, investigating the morphology of neuronal synapses during and after exocytosis. Özge Demet is a Postdoc in the lab of Prof. Dr Tobias Moser at Institute for Auditory Neuroscience, Göttingen, Germany, investigating synaptic physiology.

Jana Kroll

How would you explain the main findings of your paper in lay terms?

J.K. and Ö.D.Ö.: We were studying synaptic transmission in inner hair cells of the mouse cochlea. These cells show remarkably high rates of vesicle release and turnover. In previous studies, key neuronal fusion proteins such as the SNARE protein VAMP2 were shown to not be relevant for exocytosis at the so-called ribbon synapses in inner hair cells. Instead, a unique (and not fully understood) release machinery coordinates vesicle cycling. We were eager to test if the recycling of vesicles is likewise adapted in inner hair cells and thus characterized mouse mutants lacking the clathrin adaptor protein AP180. Performing electrophysiological, light microscopy and electron microscopy experiments, we could show that AP180 indeed plays a key role in the reformation of synaptic vesicles. Inner hair cells from AP180-knockout mice lost more than 50% of their cytosolic vesicle content compared to wild type, which is a much more drastic effect than what was previously described for neuronal synapses. Beyond that, we also noticed a problem in sustained exocytosis that we attributed to a defect in release site clearance. This finding was surprising for us because in neurons, AP180 is linked to the specific sorting of VAMP2 post-fusion.

Özge Demet Özçete

Were there any specific challenges associated with this project? If so, how did you overcome them?

J.K. and Ö.D.Ö.: Not only in this study, but more generally, working with inner hair cells is challenging and requires some training. It starts with the acute preparation of organs of Corti from mice shortly after the onset of hearing (14–17 days after birth). This process is fiddly as hair cells easily get damaged when the experimenter is too harsh or too slow.

Then, performing the actual experiments is also tricky; within the organ of Corti, there is only one row of inner hair cells surrounded by different supporting cells.

J.K.: For electron microscopy experiments, we first need to ‘find’ this one row of inner hair cells within the embedded sample. For that, we made sections, checked them at the electron microscope, cut a bit deeper, made new sections, checked again,… always keeping in mind that if one cuts a few micrometers too deep, the sample is gone. When we finally reached the basal half of hair cells, where the ribbon synapses are located, we usually screened lots of sections until we found a synapse every once in a while.

ÖDÖ: For patch-clamp experiments, after acutely dissecting the organ of Corti, supporting cells have to be carefully removed by a suction pipette in order to reach the one row of inner hair cells. This ‘cleaning’ step can be quite tricky as, if it is not carefully done, it can damage the inner hair cells. Health of the inner hair cells has a critical importance for successful electrophysiological recordings. Furthermore, in this study, we measured exocytosis and endocytosis by changes in membrane capacitance, which require long-lasting stable recordings.

Have you had any significant mentors who have helped you beyond supervision in the lab? How was their guidance special?

J.K. and Ö.D.Ö.: Apart from our supervisors who have helped and supported us whenever and wherever required, we see the whole Institute for Auditory Neuroscience as a mentor, a guide, and a fantastic and collaborative working place.

What motivated you to pursue a career in science, and what have been the most interesting moments on the path that led you to where you are now?

J.K.: At school, it was clear for me that I would be a researcher one day. I studied molecular medicine in Göttingen and was particularly interested in immunology. During my studies, I had to perform lab rotations in different fields, also including neuroscience. At the beginning of my lab rotation with Carolin Wichmann, I said to her “To be honest, I am not that interested in neuroscience and will probably just stay for this one lab rotation.” At the end of my lab rotation, after she introduced me to electron microscopy and to these special ribbon synapses, I asked if I could do my masters thesis with her and finally also graduated in neuroscience with Carolin as my co-supervisor.

ÖDÖ: I have been interested in science since school. I studied molecular biology and genetics in Istanbul (Turkey). I am dedicated to challenging myself, and pursued a PhD in Germany. What led me to neuroscience was the passionate journal clubs and physiology classes we had during my studies.

What's next for you?

J.K.: I started my first postdoc in the lab of Christian Rosenmund in the Institute of Neurophysiology at the Charité Universitätsmedizin Berlin. What changed was my model system and my research focus. I switched from inner hair cells to neurons and from endocytosis to exocytosis. I will keep doing electron microscopy and hopefully even expand my methodological repertoire. In a couple of years, I would like to become a group leader.

Ö.D.Ö: I have recently finished my PhD. I currently work as a postdoc in the same lab (Institute for Auditory Neuroscience) to finalize other projects I have been working on. I plan to move to another research topic and lab in synaptic physiology but to work on another system.

Tell us something interesting about yourself that wouldn't be on your CV

J.K.: I started to play chess when I was 12 years old – not very successful but good enough keep doing it. I am a persistent and strategic player, so I don't mind spending hours to think about solutions for one problem. I guess it is not just a coincidence that I have become passionate about electron microscopy(!).

Jana Kroll's contact details: Institute of Neurophysiology, Charité Universitätsmedizin Berlin.

E-mail: jana.kroll@charite.de

Özge Demet Özçete's contact details: Institute for Auditory Neuroscience, Göttingen, Germany.

E-mail: oezge.oezcete@med.uni-goettingen.de

Reference

Kroll
,
J.
,
Özçete
,
D. Ö.
,
Jung
,
S.
,
Maritzen
,
T.
,
Milosevic
,
I.
,
Wichmann
,
C.
and
Moser
,
T.
(
2020
).
AP180 promotes release site clearance and clathrin-dependent vesicle reformation in mouse cochlear inner hair cells
.
J. Cell Sci.
132
,
236737
.