Cilia are motile
protrusions from a myriad of specialized cells. Their role in
most systems is biological transport, from propulsion of
protozoa in water to removal of foreign matter in mammalian
trachea and ingestion assistance in frog’s esophagus.
Mucus propelling
cilia are 5-7
mm
long, 0.2 mm
thick, densely packed (~ 1
mm
interciliary spacing) and continuously beating. |

SEM of mucus propelling cilia |

Commonly
accepted beat cycle of a cilium. Positions 1-9 : fast effective
stroke. Positions 9 – 12: slow recovery stroke (on an inclined
plane) |

An AFM tip is
dipped in a field of beating cilia. Cilia hitting the tip are
causing it to rock, and its motion is detected by the laser beam
focused on its back. |
|
The experimental setup consists of a
flat-scanner AFM, leaving a
clear optical path for two optical microscopes. Simultaneously
with the AFM measurement, light is shone on the ciliary cells
through one objective, and collected through the other
objective, after passing through the beating cilia. The
modulated light is converted with a PMT and sampled with a
digital card. |
 |
The force
sensed by the AFM increases with the decrease of probe-sample
separation.
The deeper the
probe penetrates the more cilia hit it. The curve is starting to
bend up at around 5
mm,
which correlates nicely with the known length of the cilia.
Our
measurements show:
Applied
force per cilium
710 ± 230
pN |
 |
Simultaneous
optical measurement assures that no mechanical stimulation is
induced by the AFM probe. The frequency before and after AFM
probe approach are identical. Stimulation of beat frequency with
extracellular ATP causes identical frequency increase in both
signals. |
 |
AFM
measurement showing the amplitude and frequency of
the AFM tip, as the cilia are stimulated with extracellular ATP.
The
relationship
frequency-amplitude is linear:
the log-log plot has a slope of 1. This indicates that the
length of the effective stroke arc remains unchanged.
 |

Optical Measurement |

AFM measurement |
During a stimulation event, the frequency is increasing as
reported by both optical and AFM signal. The amplitude of the
AFM signal is increasing, while that of the optical signal is
decreasing. This indicates that while the arc length of the
effective stroke does not change, the plane on which the
recovery stroke is performed, becomes more perpendicular to the
plane of the cell membrane |
A field of beating cilia and the AFM tip, as viewed through the
optical microscope.
(click the image to play) |