Annals of Botany 84 : 501–506, 1999
Article No. anbo.1999.0943, available online at http :\\www.idealibrary.com on

Mechanism of Anther Dehiscence in Rice (Oryza sativa L.)
T. MATSUI*†, K. O M A S A‡ and T. H O R I E§
* Experimental Farm, Kyoto Uniersity, 12-1 Hatchonawate, Takatsuki, Osaka 569-0096, Japan ; ‡ Graduate
School of Agriculture and Life Sciences, The Uniersity of Tokyo, Tokyo 113-8657, Japan and § Graduate Schoolof Agriculture, Kyoto Uniersity, Kyoto 606-8502, Japan
Received : 5 April 1999

Returned for revision : 24 May 1999

Accepted : 23 June 1999

This paper presents a new explanation of the mechanism of anther dehiscence in rice during the period from floret
opening to pollen dispersal. The theca dehisced on the stomium in the apical part and the anther wall in the basal
part of the large locule. Comparison of the anther dehiscence process under various air humidity conditions showedthat the process, until the splitting at the apical and basal parts, was a moisture-requiring process whereas thewidening of the splits in both parts was a desiccatory process. Observation of the anther transverse section, revealed
the marked development of the U-shaped thick cell wall in the endothecium adjacent to these two splits. From theseobservations, the anther dehiscence mechanism may be explained as follows. At the time of anthesis, pollen grains
swell rapidly in response to the floret opening and cause the theca to bulge, rupturing the septum. The pollen pressure
combined with the inward bending of the locule walls adjacent to the stomium causes splitting of the stomium in theapical part of the theca. At the same time, the septum rupture extends to the bottom of the large locule supportedby the pollen pressure. After these processes, the locule walls adjacent to both splits straighten probably due to theirwater loss. This straightening widens the splits and the swollen pollen grains overflow from the widened splits.
# 1999 Annals of Botany Company
Key words : Anther dehiscence, Oryza satia L., pollen grain swelling, rice, septum, stomium, theca.

INTRODUCTION
Pollination is an indispensable process for the fructificationof grain crops. Thus, poor dehiscence of anthers causes a
decreased yield of grain crops. In Asian rice, (Oryza satiaL.) which is a major staple food in East Asia, cool weather( 20 mC) during the booting stage and high temperature( 35 mC) at flowering lowers the yield (Horie, Yajima andNakagawa, 1992 ; Horie et al., 1996), mainly through poor
anther dehiscence (Shimazaki et al., 1964 ; Ito et al., 1970 ;Satake and Yoshida, 1978 ; Matsui et al., 1997 a ; Matsui,
Omasa and Horie, 1997 b). Anther dehiscence is the mostsusceptible event to temperature stresses in rice production.
Despite the importance of the anther dehiscence in rice,
little attention has been paid to this process. Although, as inother anemophilous plants, it has generally been believedthat the rice anthers open as a result of the outward bendingof the locule walls caused by their desiccation (Hoshikawa,1993), there are several lines of evidence against this theory.
Tsuboi (1961) found that floret opening was hastened in rice
by rubbing the panicle with ones hands and that the anthersin the florets thus opened at 3 h before the expected
flowering time and could not dehisce. This poor dehiscenceindicates that the anther has a low ability to dehisce until 3 h
before flowering time, thus suggesting that the mechanismof anther dehiscence is not as simple as the above theory.

† Corresponding author. Fax j81 726 83 1532.

0305-7364\99\100501j06 $30.00\0Moreover, as evaporation from the anther is increased at ahigh temperature and the desiccation of the locule wall is
accelerated, the disturbance of anther dehiscence by hightemperature at the time of flowering observed by Satake andYoshida (1978), means that the desiccation of the locule wall
may not be the main driving force to open the anthers asshown in Liliaceae (Keijzer, 1987, 1999).
In connection with the above issue, Matsui, Omasa andHorie (1999) found a rapid swelling of pollen grains in
response to floret opening in rice, and pointed out its
importance in anther dehiscence. The purpose of the present
article is to clarify the mechanism of anther dehiscenceduring the period from floret opening to pollen shedding.
For this purpose, we observed the structure of the riceanther and the process of anther dehiscence, and examinedthe effect of air humidity on this process.

MATERIALS AND METHODSTranserse section and process of anther dehiscence
Paddy rice (Oryza satia L. ‘ Nipponbare ’) plants weregrown in submerged 4 l pots under outdoor conditions.
Intact anthers were collected and fixed in FAA on the day
before flowering (the day of flowering was judged from theposition on the panicles). The fixed anthers were dehydrated
with a methanol-butanol series, and embedded in paraffin.
Ten-micron-thick transverse sections were cut and stainedwith toluidine blue-O.
# 1999 Annals of Botany Company

502

Matsui et al.—Mechanism of Anther Dehiscence in Rice

Air humidity (% R.H.)

Air temperature (°C)

Onset of flowering

RESULTS

35

Structure of the anther

30
25
20
0

6

12

18

24

0

6

12

18

24

6

12
Time of day

18

24

100
80
60

Process of anther dehiscence40
30

Illumination (klx)

The rice anther consists of two thecae, bound together
with connective tissue (Figs 2 and 3). Each theca has a long
(longer at the base) and a short locule. These two locules
are linked with small epidermal cells (stomium) and septum(Fig. 3 A, B, C). Although a cavity for the dehiscence forms
in the centre of the septum, some of the septum cells still
persist (Fig. 3 C), keeping the locules closed, supported by
the deteriorated tapeta. The U-shaped cell-wall thickening
in the endothecium develops better in the apical part
especially adjacent to the stomium, and in the basal part ofthe locule than in the other parts (Fig. 3 A, B, D).

20
10

0

F. 1. The phytotron conditions. Wind velocity was fixed at 0n5 ms−".

Apart from this, intact florets of the paddy rice(‘ Akitakomachi ’) grown under the same condition werecollected just before flowering time to observe the antherdehiscence process. After removal of the lemma, the processof anther dehiscence was observed under a stereo microscope. Observation began within 30 sec from the samplingof the anthers.

After removal of the lemma, the process of antherdehiscence began immediately. The first detectable change
relating to anther dehiscence was the rapid outward
emergence of the whole stomium (Fig. 4 A, B). The alreadypartly disappeared septum (Fig. 3 A, B, C), which had kept
the two adjacent locules separated, became completely
ruptured, thus dissociating the stomium from the connective
tissue. At the same time, the locule wall at the base of thelarge locule ruptured along the extension line of the ruptured
septum (Fig. 4 C). Subsequently, the stomium ruptured at
the apical part of the theca (Fig. 4 D). Within a few minutes,
the slits in the stomium and the locule wall began to widen
and pollen grains overflowed from both widened slits (Fig.
4 E).
From the above observation, the process of antherdehiscence in rice can be divided into two steps : (1) ruptureof the whole septum, and splitting of the locule wall at thebasal part of the large locule (stomium) and splitting of
the stomium at the apical part of thecae (step 1) ; (2) the
widening of the slits at the apical part of the theca and the

Effect of air humidity on anther dehiscence
Oryza satia L. (‘ Akihikari ’) plants were grown in agreenhouse (25 mC, 14-h day length supplemented withartificial light) in the submerged 4 l pots, and acclimatized
to the conditions shown in Fig. 1 for more than 3 d at theflowering stage. The florets expected to flower on a givenday (the day of flowering was judged from the position on
the panicle) were sampled at noon (about 1 h before
maximum flowering) and artificially opened by removal oflemma. The opened florets were immediately subjected tovarious air humidity conditions (100 %, 60 % and 20 %R.H.) at 24 mC. After 30 min, the percentages of dehisced
stomiums and septums were calculated. The anthers in theflorets subjected to 100 % R.H. were then exposed to open
air (60 % R.H., 24 mC) and the process of anther dehiscence
was observed. Five florets were used for each treatment andthe experiment was repeated twice.

F. 2. Exterior of a rice anther.

Matsui et al.—Mechanism of Anther Dehiscence in Rice503

F. 3. The transverse sections of rice anther. (A) apical part of anther (L, locule ; P, pollen grains ; arrowheads, well developed cell wall thickening
in endothecium ; bar l 100 µm) ; (B) middle part of anther (L, locule ; P, pollen grains ; bar l 100 µm) ; (C) magnification of cavity for dehiscencein middle part of anther (L, locule ; P, pollen grains ; (C) cavity for dehiscence ; arrowhead, stomium ; bar l 25 µm) ; (D) Basal part of long locule(L, locule ; P, pollen grains ; arrowheads, well developed cell wall thickening in endothecium ; bar l 50 µm).

basal part of the large locule, and the overflow of the pollen
grains from both of the widened slits (step 2).
Figure 5 shows schematically the shape of the antherwalls during both steps. At step 1, the whole locule wall (AB)became bulged at the middle part of the theca. At the apicalpart of the theca, the part of the locule wall which faced theother locule (BC) remained bent inward while the otherparts of the locule wall (AB) became bulged. At the basal
part of the large locule, the same part of the locule wall (BC)remained bent inward while the other parts (AB) becamebulged. At step 2, the parts of the locule wall which had
remained bent inward at step 1 (BC), at the apical and thebasal parts, straightened. The shape of the locule wall at
the middle part remained unchanged at this step. The partsof the locule wall that were bent inward at step 1 andstraightened at step 2 (BCs), coincided with the parts of thelocule wall having the well-developed U-shaped cell wall
thickening.

Effect of air humidity on the process of anther dehiscence
Only 46 % of the septa were ruptured at a R.H. of 20 %,and the percentage of ruptured septa increased with the504

Matsui et al.—Mechanism of Anther Dehiscence in Rice

F. 4. Photographs showing the process of anther dehiscence. (A) just after removal of the lemma ; (B) rising of stomium (arrowhead) ; (C) split oflocule wall at the basal part (arrowhead) ; (D) split of stomium at the apical part (arrowhead) ; (E) overflow of pollen grains from both the widened
slits at the apical and basal parts (arrowheads).

increase in air humidity (Table 1). At a R.H. of 100 %, allsepta were ruptured. Moreover, 80 % of stomiums weresplit at the apical part of the thecae and 100 % of the locule
walls at the basal part of the large locules were split along
the extension lines of the ruptured septa. However, the slits
ruptured at a R.H. of 100 % were narrow and most of thepollen grains were held in the thecae. After subsequent
desiccation in the open air (60 % R.H.), the remaining
unruptured stomiums split, and both of the slits on stomiumsat the apical and basal parts widened (data not shown).
Stomiums on the thecae with unruptured septa did not open
under any humidity conditions.
DISCUSSION
Hoshikawa (1993) reported that the outward bending of
locule walls in rice caused by desiccation, causes ruptureof the septum and splits the stomium. In contrast, Cheng,
Greyson and Walden (1979, 1986) suggested that in maize,
rupture of the septum does not involve endothecial fibrouswall thickening which generates the outward force on the
locule wall by desiccation, and assumed that the shorteningof the outer epidermal wall, from microspore stage to thetime of anther dehiscence, generates the tensile force
responsible for the separation of the stomium fromthe underlying connective tissue. Our experimental data,
however, suggested that desiccation is unnecessary forthe rupture of septum ; rather, the desiccation disturbs the
rupture, in line with the findings of Keijzer (1987). Thus
septum rupture is a moisture-requiring process.

On the other hand, Matsui et al. (1999) found that pollen
grains swell rapidly in response to floret opening and thatthe swelling is indispensable for anther dehiscence. Moreover, they compared the rupture of the septum in the anthersegments containing pollen with that in the anther segments
lacking pollen under the submerged condition, and showedthat the swelling of pollen in water causes the rupture of the
septum. They infer from these data that the rapid swellingis the driving force of septum rupture in rice. The present
results also indicate that septum rupture is a moisturerequiring process, in agreement with their hypothesis.
Keijzer (1987) showed that enzymatic breakdown of cellwall material and breaking tapetal membranes by the pollenpressure lead to the opening of the septum in Gasteria
errucosa. Moreover, Keijzer, Leferink-ten Klooster andReinders (1996) reported that in maize, the formation of thedehiscent cavity is a lytical-enzymatical process preceding
the dehiscence and that such an enzymatic cell wall
breakdown is found even in the stomium. In the experimentof artificial water-induced septum rupture by Matsui et al.
(1999), the septum which had not dehisced by the artificial
opening of the floret, probably because of the low ability ofpollen grains to swell, dehisced by soaking in water. Theirdata indicate that the rupture does not depend on thechange in the physical character of the tissue but on theswelling ability of the pollen grains. The rapid swelling ofpollen grains would be a driving force to rupture theseptum, weakened by the lytical-enzymatical process.
The stomium split only in the apical part of the theca at
step 1. At the middle part, the whole theca bulged and theMatsui et al.—Mechanism of Anther Dehiscence in Rice

F. 5. The shape of locule walls in anther the dehiscence processes.
Dotted lines indicate original position of locule walls. Ll, large locule ;Sl, small locule ; C, cavity for dehiscence ; Ct, connective tissue.
Step 1 : Rupture of whole septum, split of stomium in the apical part andsplit of locule wall in the basal part. Step 2 : widening of the slits in thestomium in the apical part and in the locule wall in the basal part.

T     1. Effect of air humidity on theca opening process

Relative
humidity (%)

Ruptured
septum (%)

20
60
100

46n3a
74n1a,b100n0b

Ruptured
Ruptured locule
stomium in thewall in theapical part ofbasal part ofthecae (%)
large locule (%)
46n3a
74n1a
80n0a

46n3a
74n1a,b100n0bWithin columns, means with same letters are not significantly
different (P 0n01).

slit of the locule wall on the stomium stretched, but thestomium did not split. This indicates that the pollen pressurealone could not split the stomium. From observation of thelocule wall at the split, we suppose that the bulge of thetheca caused by pollen pressure combined with theremaining inward bending of the locule walls adjacent to thestomium, stretches the stomium more severely in the apicalparts than in the middle part and causes it to split. In ourobservation of the transverse-section of the anther, thelocule walls adjacent to the splitting stomium showed themarked U-shaped thickening in their endothecium. Since
locule walls with such well developed U-shaped thickening505

in their endothecium bend outward by desiccation andinward by water uptake (Keijzer, 1987), the above theory
that the inward bending of the locule walls adjacent to thestomium helps the stomium to split, is in agreement with thepresent data indicating that the desiccation of the locule
wall is unnecessary until the stomium splits. The remaining
inward bending will be maintained by turgor in the locule
wall. Keijzer (1987) found inward bending of the locule wallin G. errucosa disrupts the stomium with the help of theepidermal cells adjacent to the stomium acting as a fulcrum.
Such assistance of the epidermis was not found in ourobservation ; the locule walls adjacent to the splitting
septum remained separate.
At step 1, the locule wall in the basal part of the large
locule splits along the extension line of the ruptured septum.
This rupture was observed at a R.H. of 100 %, indicatingthat it is not a desiccatory process. Since a theca consists of
only one locule in its basal part, the locule wall would be
ruptured by pollen pressure as in the case of the septum
which kept the locules closed in the upper part.
At step 2, the locule walls adjacent to the longitudinal slits
at both apical and basal parts became unbent and the slits
widened. This is a desiccatory process because the locule
walls remained bent under 100 % R.H. even after rupture of
the septum and straighten with subsequent desiccation. The
desiccation will straighten the parts of the locule wall having
well-developed U-shaped thickening in the endothecium by
the combination of shrinkage of the tangential outer walland fixed inside circumference as observed in other plants(Keijzer, 1987).
In conclusion, the swelling of pollen grains is the driving
force for the final rupture of the whole septum and for the
splitting of the stomium in the apical part and the locule
wall in the basal part, and U-shaped cell wall thickening is
the key structure for the splitting of the stomium in theapical part and for widening of splits both in the apical andbasal parts. However, the outward force caused bydesiccation of the locule wall is not sufficient to rupture theseptum and stomium.
The process of anther dehiscence in rice can be describedas follows : (1) at the beginning of anthesis, pollen grains
swell rapidly in response to the floret opening and theincreased pollen pressure causes the theca to bulge, rupturing
the septum ; (2) at this stage, pollen pressure, combined with
the inward bending of the locule walls adjacent to stomium,causes the stomium to split at the apical part of the theca ;(3) at the same time, the septum split runs to the bottom of
the large locule along the extension line of the septum
supported by the pollen pressure ; (4) after these processes,
the locule walls adjacent to the splits in the basal part of thelarge locule and the apical part of the thecae straighten due
to their water loss ; and (5) such straightening of the locule
wall widens the splits in the apical and basal parts, and theswollen pollen grains overflow from both widened splits.

A C K N O W L E D G E M E N TS
We thank Mr M. Mineyama for his technical assistance.
This research was supported in part by a Grant-in-Aid for506

Matsui et al.—Mechanism of Anther Dehiscence in Rice

Scientific Research (No. 09760016) from the Ministry of
Education, Science and Culture, Japan.
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