CAPITULO III: ANÁLISIS DEL MERCADO
5.1.5. valuación Económica del Proyecto
5.1.5.1. Aspectos Generales
100um
lOOum
Figure 3. Diagrammatic representation of org an o g en esis during the embryonic development of the plaice.
(a) Formation of organs within the 8°C and 12°C embryos expressed in days post fertilisation.
(b) Organ formation at 8°C and 12°C shown as a proportion of total development time.
Development was a s s e s s e d by examining transverse and longitudinal wax sections of embryos sam pled daily between fertilisation and hatching.
Horizontal bars illustrate myogenesis, myofibrillargenesis and formation of the notochord, spinal cord and gut in the anterior myotomes. Formation of the eyes, neurocoel, ventral vessel (presumptive dorsal aorta), dorsal (D) and ventral (V) finfolds, pronephros and pectoral fins are indicated by arrows. Muscle fibre formation was divided into three sta g es; (1) formation and division of p re myoblast cells; (2) alignment and fusion of myoblasts to form myotubes; (3) appearance of muscle fibres containing discrete myofibrils. Progressive changes in the appearance (maturity) of the notochord, spinal cord, gut and muscle fibres are illustrated by the density of pattern within the horizontal bars.
(a)
1 2 °C y 't//>/''/,'//////,'/,\ Spinal cord i N eurocoel Ventral vessel f 'f '//y///r//////,'/////\ N otochord Eyes i I Pronephros D/V finfolds ] Myofibrillargenesis Pectoral fins____________y Superficial fibres
. ; .•. Muscle fibres Pre-m yoblasts M yotube formation
8 ° C y "/ '/ '> '///'/r///^//////////////JJ\ Spinal cord A Neurocoel Ventral vessel f > / / / / ! ' I / ' ; ' ; I > / I I / / , / / ' / / / ' / ' , Notochord Eyes Pronephros D/V finfolds Pectoral fins Pre-myoblasfs / J J M yofibrillargenesis ' ' . . . . T Superficial ftores... . ' I : : \ Mus cl e fibres Myotutje formation 10
D ays post fertilisation
15 (b) * n * n 1 1 i • 11 • II t'ï _ ' / / / / / / / / , ' / , ' , V / / / J G u t '/ ' / ' / ' / ' / ItllllllHllllllllls p m a i c o r d. N e u r o c o e l A y V e n tra l v e s s e l ^ ' / ' / : / / / / / / / / / / / / / / / A111 11 11 I 1 1 11 1 11 11 N o to c h o r d i 1 E y e s P r o n e p h r o s
D /V finfolds j M y o fib rilla rg e n e s is P e c to r a l fins y S u p e rfic ia l fib re s
• It' r' • M u s c le fib re s P r e - m y o b la s ts M y o tu b e f o rm a tio n / / / / / / / / / ' / / / ! I I ! / / / / , / / / / 1 1 1 1 / / i * I f 1 I ^ A ^/ / / / / / / / / / / / / , ! G u t / : ' / '/ c o r d N e u r o c o e l A y V e n tra l v e s s e l * i ' t ^ t ' f ' / ^ / * / / / • ; II t II 1 • ! 1 II ! 1 t ' \ . ,N o to c h o r d i i E y e s P r o n e p h r o s
D /V fin fo ld s J M y o fib rilla rg e n e sis P e c to r a l fin s f S u p e rfic ia l fib re s
rEZj M u s c le fib re s P r e - m y o b la s ts M y o tu b e fo rm a tio n
12°C
8°C
25 50
Percentage hatch time
100um
Figure 4. Plaice embryo reared at 12°C, five days a f t e r fertilisation. Myoblasts are arranged in rows, parallel to the notochord. Longitudinal wax section stained with Haematoxylin and eosin.
after fertilisation it was possible to distinguish the
superficial muscle fibres from the inner fibres in the embryos maintained at 12°C (Fig. 3a).
Organogenesis was slower at 8°C; at 6 days post
fertilisation the 8°C embryos reached the same developmental stage as the 12°C embryos had reached at 4 days post
fertilisation (Fig. 3a). The order in which the various organs and tissues formed in the embryos was the same at both incubation temperatures. At both 8°C and 12°C the different organs differentiated from the embryonic mesoderm at
equivalent points between fertilisation and hatching and were present in the developing embryo for a very similar proportion of the period between fertilisation and hatching (Fig. 3b).
The information obtained from the wax sections can only be used to give an estimated time for the events comprising muscle fibre differentiation in plaice embryos. The
magnifications provided by the light microscope (x20-1000) are too low to distinguish the initial stages of
myofibrillargenesis and myotube formation. T.E.M data is required for a more accurate timetable. Plaice eggs were fixed and embedded for examination with the T.E.M, but the chorion, even when punctured was found to completely block the passage of osmium and unfortunately the developing
embryos were too small to successfully survive the embedding process when dissected free from the egg capsule.
Effect of incubation temperature on embryonic somite d e v e l o p m e n t
Development time, defined as the period between
fertilisation and 75% of the embryos hatching, was 456 hours at 5°C and 210 hours at 12°C. The head and eye capsules had formed when the embryos were examined after 22% (104 hours) development time at 5°C and 23% (46 hours)
development time at 12°C but distinct somite boundaries were
not present. Somites could first be counted after 25% (114 hours) development time in embryos reared at 5°C, when 17 ± 1.1 (n = 8) somites were present (Fig. 5). At 12®C, 14 ± 1.5 (n = 8) somites were visible after 31% (65 hours) development time (Fig. 5). This suggests that formation of the initial
somites occurs relatively later in development at high than at low temperatures. At any given stage of development embryos reared at 5°C had significantly more somites than the embryos maintained at 12°C (p < 0.05). For example, after 35%
development time 5°C embryos possessed 33 ± 0.9 somites while 12°C embryos, after 36% development time, had 26 + 1.2 somites (n = 8). After 70% and 69% development time
respectively, 5°C embryos had 52 ± 1.1 somites and 12°C embryos 46 ± 1.0 somites (n = 8). At both stages the
differences in somite numbers were statistically significant (p < 0.05). Spontaneous contractions of the anterior somites and a regular heart beat were observed after 47% of
development time at both temperatures, corresponding to the 50 somite stage at 5°C and the 44 somite stage at 12°C (Fig. 5). Somitogenesis proceeded more rapidly at both
temperatures before the embryos began moving (Fig. 5a and b). At 5°C, somites were added to the tail at a rate of 0.35
somites/hour. Prior to spontaneous movement in the 12°C embryos the rate of somite addition was 0.8 somites/hour (Fig, 5a).
When the embryos hatched the total length of the larvae reared at 5°C was 6.61 ± 0.36 mm (n = 10) while the 12°C larvae measured 6.07 ± 0.22 mm (n = 10), a 9% difference (p < 0.05). Newly hatched 5°C larvae possessed 61 ± 1 . 5 body somites (n = 8), 9% more than the 56 ± 1.3 (n = 8) somites of 12°C larvae at hatching (p < 0.05).
Figure 5.
(a) The rate of somite development in embryos maintained at 5°C and 12°C.
(b) Somite development in embryos reared at 5°C and 12°C expressed a s a percentage of the total embryonic period (defined as the period between fertilisation and 75% of the embryos hatching).
Values represent mean ± standard deviation (n = 8 embryos/temperature). Note some of the error bars fall within the sym bols. Arrows indicate the start of spontaneous movement by the embryos.