Biomedical Engineering Reference
In-Depth Information
The cercal sensory system in the cricket detects the direction, frequency, and
velocity of air currents with great accuracy and precision. The receptor organs of this
system consist of a pair of antenna-like appendages called cerci at the rear of cricket
abdomen. Each cercus is covered with approximately 500 fi liform hairs, each of
which is innervated by a single mechanoreceptor neuron. The receptor neuron is
tuned to air currents from a particular direction and exhibits a change in its fi ring rate
in response to stimuli over the entire 360° range of stimulus directions (Landolfa and
Miller
1995
). Optical imaging of a population of the mechanosensory afferents
stained with the AM ester of a Ca
2+
indicator demonstrated that the direction of the
air currents is represented by specifi c spatial patterns in the ensemble activities of the
afferents (Ogawa et al.
2006
). Identifi ed giant interneurons (GIs) receive direct excit-
atory synaptic inputs from the mechanosensory afferents. The GIs are activated by
air currents and also display differential sensitivity to variations in air-current direc-
tion (Jacobs et al.
1986
; Miller et al.
1991
; Theunissen et al.
1996
). The directional
tuning curves of the GIs are well described by a cosine function; that is, the GIs
encode information about the stimulus direction proportional to their spiking activ-
ity. To describe decoding algorithm of the directional information from the popula-
tion activities of sensory afferents to individual GIs, we measured the pre- and
postsynaptic local Ca
2+
responses to the air-current stimuli, on each GI's dendrite.
Two kinds of Ca
2+
indicators with different fl uorescent wavelengths were loaded
to pre- and postsynaptic neurons, respectively. The sensory afferent fi bers were
stained with AM ester of Oregon Green 488 BAPTA-1 (OGB-1), while cell-
impermeant Fura Red was injected into the single GI (Fig.
5.4a
). For staining the
sensory afferents, a solution containing AM ester of OGB-1 at a concentration of
0.05 % and dispersing reagent (Pluronic F-127, Invitrogen) at a concentration of
1 % was pressure injected into a cercal sensory nerve through a glass micropipette.
Twelve hours after dye injection, the axon terminals of cercal sensory neurons were
found to be stained with OGB-1. After staining of the afferents with OGB-1, 2 mM
Fura Red tetrapotassium salt was iontophoretically injected into the GI for 5 min
through a glass microelectrode, using a hyperpolarizing current of 3 nA. Fluorescent
signals were viewed with an inverted microscope (Axiovert100, Zeiss). A xenon arc
lamp (XBO 75 w, Zeiss) illumination with a stabilized power supply and 470/20
band-pass fi lter were used for excitation of the Ca
2+
indicators, OGB-1, and Fura
Red. For simultaneous measurement of pre- and postsynaptic Ca
2+
signals, a fl uo-
rescent image passing through a FT510 dichroic mirror was divided into two images
with
W-View
optics (Hamamatsu Photonics) by the following fi lter set: dichroic
590LP, emission 535/45 for the OGB-1 and 610/25 for Fura Red (Fig.
5.4b
). The
two separated images were simultaneously acquired side by side in the same frame
with a digital cooled CCD camera (ORCA-ER, Hamamatsu Photonics) attached to
the inverted microscope.
Using this optical system to separate the optical signals from OGB-1 and Fura
Red, we simultaneously measured the Ca
2+
responses to air-current stimuli in the
GI's dendrites and in the afferent axon terminals that make synaptic connections
onto that dendritic branch (Ogawa et al.
2008
). It was observed that the air-current
stimulus evoked a signifi cant decrease in the fl uorescence of Fura Red (610 nm
