QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

This Article Summary Full Text Full Text (PDF) Movies Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Iwadate, Y. Search for Related Content PubMed PubMed Citation Articles by Iwadate, Y.

Photolysis of caged calcium in cilia induces ciliary reversal in Paramecium caudatum

Yoshiaki Iwadate^*

Department of Life Science, Faculty of Integrated Arts and Sciences, The University of Tokushima, Tokushima 770-8502, Japan

View larger version (71K): [in a new window]   Fig. 1. Photolysis of caged calcium. An image of a pinhole with a diameter of 400 µm was focused onto the specimen. (A) 300–400 nm UV light was collected by a UV-transmitting objective lens to produce a small UV spot on the specimen. (B) A Paramecium caudatum cell. Approximately 15 000 cilia are present on the cell surface. The length of each cilium (approx. 10 µm) is equivalent to the diameter of the UV spot (A) in which photolysis was induced. Bar, 30 µm.

View larger version (13K): [in a new window]   Fig. 2. Schematic illustration of a cross section of the cortical region of a Paramecium cell. The cilium (c) grows from a basal body (bb) inside the cell membrane (cm). The terminal plate (tp) separates the basal body and the upper region of the cilium. The trichocyst (t) and alveolar sac (as) are located in close proximity to the basal body. The diameter of the cilium is approx. 200 nm and the length of the trichocyst, 5 µm. Following exocytosis, the released trichocyst expands 25 µm in length.

View larger version (46K): [in a new window]   Fig. 7. Cell function in response to UV application. Light was applied to the whole cilium, including both the basal body and upper region, at the anterior of a Paramecium cell loaded with NP-EGTA in Ca2+-containing medium. (A) A series of images around the UV application area (time course is indicated above each picture). UV light was applied to the area indicated as a circle in the picture marked 0.17 s. UV light was somewhat scattered by cytoplasmic particles. Arrowheads mark trichocysts released via exocytosis indicating the [Ca2+]i at the basal bodies; arrows, direction of the cilia inside or outside the area of UV application. The series shows a typical result from a total of 13 experiments. Bar, 20 µm. This series of images are indicated in Movie 4, which is slowed to 1/3 of the original speed. (B) Temporal relationship of the ciliary angle measured from A. The time course in B matches the time indicated at the top of each picture in A. The period of UV application is indicated by the black bar; triangle, the time of trichocyst exocytosis; solid line, angle of a cilium within the UV application area; dashed line, angle of a cilium outside the area of UV application.

View larger version (45K): [in a new window]   Fig. 8. Changes in cell function in response to UV application. Light was applied to the inner part of the anterior of a Paramecium cell loaded with NP-EGTA in Ca2+-containing medium. (A) A series of images around the area of UV application. The time course is indicated above each picture. UV light was applied to the area indicated by the circle in the picture marked 0.13 s. UV light was somewhat scattered by cytoplasmic particles. Contraction of the cell body in response to UV application was observed at the area of UV application. Arrowheads mark trichocysts released via exocytosis, indicating [Ca2+]i at the neighboring basal bodies; arrows, direction of cilia, whose basal bodies were inside or outside the area of trichocyst exocytosis. The series is a typical result from a total of 16 experiments. Bar, 20 µm. This series of images are indicated in Movie 5, which is slowed to 1/3 of the original speed. (B) Temporal relation of ciliary angle measured from A. The time course in B matches the time indicated at the top of each picture in A. The period of UV application is indicated by a black bar; the triangle indicates the time of trichocyst exocytosis. Solid line, angle of a cilium whose basal body was within the area of trichocyst exocytosis; dashed line, angle of a cilium whose basal body was outside the area of trichocyst exocytosis.

View larger version (10K): [in a new window]   Fig. 3. Determination of the ciliary angle. A cilium within or outside the area of UV application was selected, and the angle () between it and the line indicated by the arrowhead was measured. Approximately one half of the Paramecium cell body was captured by a suction pipette (s). The angle was recorded as positive if the cilium was positioned in a portion of the cell apical to the perpendicular line.

View larger version (79K): [in a new window]   Fig. 4. Changes in [Ca2+]i and cell functions of Paramecium cells loaded with NP-EGTA medium in response to UV light applied for 125 ms to the entire cell. [Ca2+]i was estimated by monitoring the fluorescence intensity of Calcium Green in the Paramecium cell. (A) [Ca2+]i values are expressed as PMT current. The value of PMT current was zero during UV application because the shutter in front of the PMT was closed to protect the PMT from the UV light. Arrow, UV application period. UV application caused an abrupt [Ca2+]i in Paramecia loaded with NP-EGTA medium. A typical result from four experiments is shown. Ciliary reversal and trichocyst exocytosis in response to [Ca2+]i is shown prior to (B) and following (C) UV application. Ciliary reversal took place all over the cell. Arrows, released trichocysts via exocytosis. Bar, 50 µm. A typical result from four experiments is shown.

View larger version (50K): [in a new window]   Fig. 5. Ciliary reversal in response to UV application. Light was applied to cilia at the anterior of a cell loaded with NP-EGTA in Ca2+-chelating medium. (A) A representative result in which the distance l is zero. The distance l was determined as indicated in the inset in (B). Before, prior to UV application; after, following UV application. UV light was applied to the area indicated by the circle. Arrows indicate direction of the cilia within or outside the area of UV application. Bar, 20 µm. This series of images is shown in Movie 1. (B) Relationship between the distance l from the rim of the area to which UV light was applied (circled) to the cell surface and the angle of ciliary reversal. The angle of ciliary reversal was calculated by subtracting the angle of the cilia prior to UV application from that after UV application.

View larger version (43K): [in a new window]   Fig. 6. Ciliary reversal in response to UV application. Light was applied to the cilia at the anterior of a cell loaded with NP-EGTA in Ca2+-containing medium. (A) A representative result in which the distance l (see Fig. 5) is zero. This series of images are indicated in Movie 2. (B) The distance l is 6 µm. This series of images are indicated in Movie 3. Before, prior to UV application; after, following UV application. UV light was applied to the area indicated by the circle. Arrows, direction of the cilia within or outside of the area of UV application. Bars, 20 µm. (C) Relationship between the distance l and the maximum angle of ciliary reversal. Ciliary angles before (triangles) and after (circles) UV application are plotted against l.

View larger version (18K): [in a new window]   Fig. 9. Ciliary reversal in response to the photolysis (circled) of caged calcium at various portions of the cilia. (A) Weak reversal in response to application of UV to the tip of the cilia (cf. Fig. 6B). (B) Strong reversal in response to application of UV above the basal bodies (cf. Figs 5A, 6A). (C) Restriction of initial response to UV-treated cilia when UV was applied to both basal bodies and upper ciliary region. Following trichocyst exocytosis (t), ciliary reversal took place on the entire cell surface (cf. Fig. 7). (D) No cilium reversed their direction in response to UV application at the inner portion of the cell, although trichocyst exocytosis (t) took place, indicating [Ca2+]i at the neighboring basal bodies. After trichocyst exocytosis (t), ciliary reversal occurred on the entire cell surface (cf. Fig. 8).