First published online August 25, 2003
The retina of Manduca sexta: rhodopsin expression, the mosaic of green-, blue- and UV-sensitive photoreceptors, and regional specialization
Richard H. White1,*,
Huihong Xu2,
Thomas A. Münch3,
Ruth R. Bennett1 and
Erin A. Grable1
1 Department of Biology, University of Massachusetts Boston, 100 Morrissey
Blvd, Boston, MA 02125-3393, USA,
2 MGH Cancer Center, Harvard Medical School, 13th Street, Charlestown, MA
02129, USA
3 Helen Wills Neuroscience Institute, University of California Berkeley, 145
Life Sciences Addition, Berkeley, CA 94720, USA
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Fig. 1. Illustration for method of measuring rhabdomere volumes. (A) Profiles (see
Fig. 5A) of two adjacent rows
of retinulae traced from electron micrographs of a tangential section
extending from the retinal surface to the basement membrane. (B) Inferred
plane of section projected across a light micrograph of a longitudinally
sectioned retina.
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Fig. 2. Retina of Manduca sexta. (A) Dorsal aspect of an unfixed retina
dissected free of the cornea and clear zone. Anterior is to the right. The
blue patch is the dorsal rim area. Scale bar, 0.5 mm. (B) Freehand vertical
section of a glutaraldehyde-fixed retina photographed in reflected light.
Dorsal is at the top, distal to the left. The highly reflective tracheoles (t)
extend almost to the distal surface (ds) of the retina, except in the dorsal
rim area (dr). Scale bar, 100 µm. (C) Retinulae dissected from the center
of a fixed retina photographed in transmitted light: tracheoles are opaque,
extending across the retina to just below the distal tips of the retinulae.
bm, basement membrane beneath the retina. Scale bar, 10 µm. (D) Dorsal rim
retinulae. Tracheoles envelop only the proximal ends of the retinulae. Scale
bar, 10 µm.
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Fig. 3. Rhabdomere volumes of receptor cells near the center of the retina. (A)
Light micrograph of longitudinal section showing two retinulae. bm, basement
membrane; ne, nuclei of elongate photoreceptors; np, nucleus of small proximal
receptor; r, rhabdom; t, tracheoles; te, tracheole ends. (B) Rhabdomere
volumes, calculated from 21 virtual sections (see Materials and methods and
Fig. 1A) of photoreceptors in
relation to depth of retina. Each rectangle represents a virtual section: its
vertical side represents the section thickness; the horizontal side represents
the area of the rhabdomere. The calculated rhabdomere volume for each cell
type is indicated.
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Fig. 4. Comparison of dorsal rim retinulae with nearby retinulae of the yellow
retina in the same section stained with anti-P357 and photographed with phase
interference optics to enhance images of rhabdoms. (A) Dorsal rim retinulae
with typical butterfly-shaped rhabdoms. Note the absence of surrounding
tracheoles. The narrow ventral dv cell is stained in the retinula on
the far right. (B) Retinulae surrounded by tracheoles (t) from an adjacent
region of the yellow retina. Two dv cells are stained in the retinula
on the far left; only one cell is stained in the other retinulae. Scale bar, 1
µm.
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Fig. 5. Organization of retinulae from the yellow retina and dorsal rim. (A)
Tracings of electron micrographs from sectioned retinulae near the center of
the retina, showing the component photoreceptors and their rhabdomeres
(shaded). The dorsal-ventral axis of the retina is vertical. The depth of each
retinular profile from the surface of the retina (see
Fig. 3) is indicated.
dv, dorsal-ventral receptor cells; ap, anterior-posterior
cells; ob, oblique cells; pr, proximal cell. (B) Tracing
from electron micrograph of a retinula from the dorsal rim area. The
structures of the eight elongate receptor cells do not vary with retinal depth
(the basal pr cell has not yet been found). Only the ventral
dv cell is present. Orientations of photoreceptor microvilli are
indicated by double-headed arrows. Scale bar, 1 µm.
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Fig. 6. Proteins from Manduca retina. (A) SDS-PAGE electrophoresis of
aliquot parts of retinal homogenate equivalent to 0.1 retina, calibrated with
molecular masses of Bio-Rad protein standards. (B,C,D) Western blots with
antisera against P520, P450 and P357, respectively, showing a major band at
approximately 37 kDa. (E) Control blot lacking primary antibodies.
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Fig. 7. Longitudinal sections from the same retina immunostained with antisera to
the three characterized Manduca opsins. (A) Anti-P520. (B) Anti-P450.
(C) Anti-P357. The dark blotches just above the basement membrane (bm) in A
are screening pigment, not stain. Scale bar, 10 µm.
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Fig. 8. Adjacent immunostained sections from the ventral domain of the retina,
approximately 70 µm from the surface of the retina. The dorsal-ventral
retinal axis is vertical. (A) Anti-P520. Rhabdomeres of ap and
ob cells are stained. (B) Anti-P357. Some dv cells are
stained. A retinula in which both dv cells are stained is marked with
an asterisk. Scale bar, 10 µm.
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Fig. 9. Adjacent immunostained sections from the ventral domain of the retina. (A)
Anti-P357. (B) Anti P-450. Asterisks mark retinulae in which one dv
cell stains for P357, the other for P450. Scale bar, 10 µm.
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Fig. 10. Retinal mosaic of blue- and UV-sensitive photoreceptors. (A) Diagram of the
Manduca retina drawn to scale, showing locations of the retinal
patches illustrated in B-E. Anterior is to the right. The equatorial division
(eq) between dorsal and ventral retinal domains is indicated, as is the dorsal
rim area (dr). Scale bar, 0.5 mm. (B,C,E) Diagrammatic representations of
retinal patches mapped from adjacent sections stained for P450 and P357,
respectively. Each hexagon represents a retinula containing potentially two
dv cells. A uniformly colored hexagon represents a retinula
containing two identical cells. A hexagon divided in half horizontally is a
retinula with unlike or missing cells. Blue represents cells expressing P450;
red represents cells expressing P357; white indicates no staining by either
antiserum in that position. (B) Domain in the dorsal retina that extends into
dorsal rim region (dr), whose margin is represented by the heavy line. Some
dorsal-ventral rows of retinulae terminate before reaching the edge of the
retina. Red lines in the dorsal rim region represent the narrow dv
cells that express P357. (C) Domain in the ventral retina. (D) Large patch at
the center of the retina for which grayscale micrographs of adjacent sections
immunostained for P450 and P357 have been superimposed and arbitrarily colored
(blue, anti-P450; red, anti-P357) in Adobe Photoshop. Scale bar, 100 µm.
(E) Equatorial sector from the adjacent sections shown in D.
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© The Company of Biologists Ltd 2003
