Journal Club 100: Live imaging show individual stereocilia have varying spontaneous calcium activities.

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Today's journal article

Hussain S, Sedlacek M, Cui R, Zhang-Hooks W, Bergles DE, Shin JB, Kindt KS, Kachar B. Spontaneous calcium transients in hair cell stereocilia. 

Why I picked this article

The cochlea is an amazing organ that is able to sense sounds as a tiny motion of fluid inside the inner ear. Sensory hair cells inside the cochlea act as sensors. Sensory hair cells have a hair-like structure called stereocilia lined up on a surface. Mechanotransduction in the cochlea depends on ion channels found at the tips of stereocilia called MET (mechanotransduction) channels. When stereocilia bend in response to sounds, triggering movement of the inner ear fluids, MET channels open, potassium and calcium ions flow into cells, and that triggers hair cells to release neurotransmitters and let auditory neurons know sounds have arrived. Therefore, the MET channel and stereocilia functions are critical for our hearing.

Scientists have studied MET channels and stereocilia by deflecting stereocilia and taking electrical measurements. But with the conventional method, it is not possible to understand what a single stereocilium is doing, compared to the whole bundle and the cell. In this research, researchers have used direct imaging of calcium ions combined with high-performance microscopy to understand the calcium ion movement in stereocilia at rest, across development and in vivo. 

Some of the research findings

Models and indicators
  • Genetically modified mice that have calcium detection proteins in hair cells either: 
    • Membrane-localized GCaMP3 (R26-lsl-mGCaMP3 crossed to Gfi1-Cre) 
    • Cytoplasmic GCaMP6f.
    • C57BL/6 background. 
    • Ages imaged ex vivo ranged from postnatal day (P) P0–P20. 
  • Genetically modified Zebrafish that have calcium detection proteins:
    • Tg(myo6b:GCaMP6s-CAAX)
    • Larvae at 2 and 5 days post-fertilization. 
    • Visualisations focused on the Lateral line neuromasts and inner-ear cristae. 
  • Imaging and preparation:
    • Mouse organs of Corti and vestibular tissues were dissected and either cultured 24–48 h or imaged acutely (P17–P20). 
    • Spinning-disk confocal (100×, NA 1.45; 488 nm excitation) captured time-lapse calcium signals focused to individual stereocilia.
    • Zebrafish imaging used a Zeiss LSM 980 with Airyscan (63× oil, NA 1.4), time-lapse acquisition every 5–16 s over 8–15 min.
Part of Fig1C. Spontaneous calcium level change as seen by different brightness (top panel, brighter means more calcium), and different timings (bottom panel, different colours represent different timings). Hussain et al. 2025

Key findings: 
  • Researchers observed that individual stereocilia have discrete spontaneous changes in calcium levels in 4-9-day-old mice without any stimulation. 
    • The transients were spatially punctate (often mid-shaft or base as well as tips), uncorrelated between neighboring stereocilia, and too small/brief to drive whole-cell depolarization.
    • The spontaneous calcium level change was observed in the later, more developed cochlea as well as in 17-day-old. (This stage is after the hearing onset, so the system is more mature to be able to start responding to external sounds, but is still not fully mature). 
    • Spontaneous calcium level change in each stereocilium was independent. 
    • Each spontaneous calcium level change seemed to be different; the propagation distance varied even within the same stereocilium. Some had a very quick rise in calcium level, while in others it took 1-5 seconds to reach the peak concentration. 
    • Similar calcium level changes were also observed in Zebrafish in vivo. 
  • The MET-channel blockers abolished the spontaneous calcium change in the stereocilia.
    • MET channel blocker dihydrostreptomycin at 1 mM concentration reduced the calcium level change. 
    • Another MET channel blocker, amiloride at 50 μM reduced the calcium level. 
  • Calcium level change starts at or near the stereocilia tip and moves towards the base.
    • The calcium level increase does seem to start at the site adjacent to the "tip-link" of the stereocilia, and correlates with tension. 
    • However, breaking tip links with BAPTA (5 mM, 15 min) did not eliminate the calcium level change in the Zebrafish model, implying either MET-complex precursors or channels outside classic tip-link sites can stochastically gate at rest.
  • Calcium level changes occurred before the stereocilia were mature. 
Overall, the research suggests that MET channels are active to certain levels at the resting state, with membrane tension boosting their likelihood to open. 

Haruna's takeaway

This is an amazing research publication! Very exciting to see the ionic charge in a single stereocilium, and be able to monitor it over time. I wonder what this means for the overall likelihood of each hair cell, which has many stereocilia; I'm sure the provided detailed information from this research will trigger more computerised modelling studies to unveil what it means for the hair cells. 

It seems to make sense to have the spontaneous calcium level changes, to enable the system to be more sensitive so that even a little bit of deflection could tip the balance of the internal calcium and activate hair cells. I also wonder if it could provide some developmental feedback (like how long the particular stereocilia is as it grows?). This was a ground-breaking research publication and a very nice pick for my 100th journal club blog article! 

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This is Haruna's 100/100 of the 100-day challenge to post a science blog article every day! I love inner ear biology & cochlear physiology.