RECENT RESEARCH ON POSTNATAL lvlltscle DEVEMPMENT I N SWINE* H. J. SWATIAND University of Guelph Guelph, Ontario, Canada

Transcription

1 86 RECENT RESEARCH ON POSTNATAL lvlltscle DEVEMPMENT I N SWINE* H J SWATIAND University of Guelph Guelph, Ontario, Canada Introduction and Objectives The published s c i e n t i f i c l i t e r a t u r e on p o s t n a t a l muscle growth i n t h e higher v e r t e b r a t e s i s very extensive b u t access t o most of t h e important items i s provided by t h e 1974 American Society of A n i m l Science Symposium and by Burleigh (1974) The present discussion provides only a l i m i t e d presentation of a small segment of t h e whole t o p i c, namely t h e results obtained by l i g h t microscopy It is hoped, however, t h a t t h e m t e r i a l covered w i l l be of general iinterest t o Meat S c i e n t i s t s s i n c e it has, as i t s d i s t a n t goal, an improvement i n our present methods of s e l e c t i n g l i v e s t o c k f o r t h e i r meat producing a b i l i t i e s This, of course, i s t h e g o a l that has been t r a d i t i o n a l l y claimed by researchers in t h e a r e a of muscle g r a s t h d a t i n g back t o t h e pioneer work of S i r John Hammond in 1932 a t t h e University of Cambridge In t h e 44 years which have elapsed since then, no method of s e l e c t i o n based on t h e h i s t o l o g i c a l parameters of muscle mass and growth has been applied i n commercial a g r i c u l t u r e This i s a very important point t h a t can hardly be ignored and it requires some explanation The f a i l u r e t o f i n d a p r a c t i c a l a p p l i c a t i o n i n a g r i c u l t u r e o r i g i n a t e s from a t l e a s t four and possibly more f a c t o r s : 1 Muscle samples must be procured e i t h e r by biopsy from p o t e n t i a l breeding s t o c k or by postmortem sampling f r o m t e s t progeny The former operat ion r e q u i r e s s k i l l e d t e c h n i c a l labor while progeny t e s t i n g may waste valuable time or incur heavy overhead expenses * 2 Intramuscular and intermuscular v a r i a t i o n in myof i b e r diameters (Johnson and Beattie, 1973) i s considerable and compounds t h e problems of c o l l e c t i n g meaningful samples 3 Hegarty and coworkers (Hegarty, 1971) have demonstrated t h a t g r e a t care is needed t o avoid introducing a r t e f a c t u a l v a r i a t i o n i n morphometric data acquired by l i g h t microscopic methods 4 F i n a l l y, and most embarrassing of a l l, it i s not e n t i r e l y c l e a r which h i s t o l o g i c a l features are desired and which w e should s e l e c t a g a i n s t For example, s e l e c t i o n i n favor of w e l l muscled a n i m l s with increased myof iber diameters m i g h t encourage t h e development of large diameter myofibers specialized f o r anaerobic metabolism which m i g h t then produce l a c t a t e very r a p i d l y postmortem Presented a t t h e 29th Annual Reciprocal Meat Conference of t h e American Meat Science Association, 1976

2 87 The obstacles which these factors create are serious, particularly those which relate to the cost of skilled labor The only new factor in the balance sheet which might facilitate the Driginal goals of using histological parameters as a basis of selection is provided by systems for image analysis by computer Although commercially available systems can interface directly with a microscope image, the problems of defining object boundaries (myofiber perimeters) are probably most easily overcome by using high contrast photomicrographs with myof iber boundaries sharply delineated with a silver stain for endomysia1 connective tissues Apart from this possibility, the objectives of the present generation of researchers should be more humble and more realistic They can be summarized as follows: 1 To understand at the cellular level of organization, the phenotypic changes which are now quite rapidly produced by intensive selection based on objective criteria and which are widely spread by art if icial insemination or geographic movement of breeding stock 2 To understand at the cellular level what is happening in the muscles of meat animals during growth t o slaughter weight In other words, the objective is a better understanding of the phylogeny and ontogeny of muscle in meat animls In 1971, Hegarty reviewed the existing literature on myofiber growth and development for the 24th Reciprocal Meat Conference at the University of Kentucky The present report attempts to describe the progress that has been made in the intervening five years of research Myofiber numbers and the work of Staun and Goldspink In his work on the histology of muscle growth in sheep, Hammond (1932, p 492) concluded that, "-in the improved mutton breeds the number of cells in each muscle has been increased" From his own data and the reports of other investigators, Hammond (1932, p 491-2) also concluded that, '(-up to birth there has been an increase in the number of cells, and that during postnatal life increase in size of the cell occurs" The major emphasis in these earlier studies seem to have been, as it is now, to understand the phenotypic changes occuring during phylogeny and ontogeny Although the concept of using myofiber numbers as a basis for selection is implicit in these early studies, there was a delay of many years before positive assertations of this potential method became common Thus, in 1972 Staun concluded a review of his work in Copenhagen, Denmrk, with the assertation that, "-the lean meat content in the pig can be improved by selecting animals with a large rider of muscle fibers in the muscle" In 1973, Stickland and Goldspink published their studies financed by the British Meat and Livestock Commission in which they outlined a methd for complete biopsy removal of a smll muscle from the pig's foot "he number of myofibers present in this small muscle (removal of which did not impair the pig's locomotion) were to be counted and this criterion

3 was to be used to determine the pig's genetic potential for meat production Given that Staun derived heritability estimates for myof iber numbers in the longissimus dorsi muscle of between 043 and 048 (Staun, 1972, table 5) and that Stickland and Goldspink (1973) found that myofiber numbers did not change during postnatal growth, the stage might be set for a dramatic improvement in selection of swine using biopsy and histological methods Real and Apparent Numbers of Myofibers Modern histology textbooks are filled with the recent discoveries of electron microscopists and older and less spectacular inf ormtion has often been edited out One casualty of this process has been the intrafascicularly terminating myofiber These myof ibers have one or both ends terminating within the belly of the muscle and, thus, they do not run from origin to insertion of the muscle (Schiefferdecker, 1891; Schafer, 1898; Ehrdeen, 1903; Huber, 1916; Le Gros Clark, 1945; Needham, 1964; Swatland and Cassens, 1972) In a large muscle they will be absent from a transverse section taken beyond their termination so that the nuniber of myofibers appearing in the transverse section (apparent number) may sometimes be less than the number of myofibers present in the whole muscle (real number) This problem is compounded by the fact that, in muscles such as the longissimus dorsi with a complex arrangement of fasciculi, it is not possible to transect all fasciculi in a single cross sectional plane of the muscle Thus, apparent number may be less than the real number when intrafascicularly terminating myofibers or whole fasciculi are omitted from the whole muscle cross section This is shown diagramtically in figure 1 Variation in Apparent/Real Fraction The validity and meaningfulness of the indirect mthods of estimating apparent myofiber numbers in meat animals have been examined by Jiminez -- et al (1965) There are several problems, however, that have not received the attention they deserve Whether or not they will restrict or invalidate the claim that meat animals can be selected on the basis of their myofiber numbers remains to be shown Problem 1 Staun (1972), Stickland and Goldspink (19731, Miller et al (1975) and Bendall and Voyle (1967) estimated the apparent number of myofibers in longissimus dorsi muscles by multiplying the cross sectional area of the whole muscle by the number of rqyofibers per unit area in transverse sections It is very difficult, however, to find myofibers that are parallel to the vertebral column in the lumbar region of meat an-ls Thus, in a whole muscle cross sectional area taken perpendicularly to the vertebral column, most myofibers are transected obliquely so that each myofiber has an enlarged cross sectional area (Swatland, 1975a)

4 LATERAL PLAN OF FASCiCULFtR ARRRNQEMENT IN 7YE PORCINE L, D MUSCLE TRANSVERSE SECTION IN SELTION LATERAL PIAN OF I

5 When t h e mean packing d e n s i t y of Inyofibers per u n i t area i s estimated i n h i s t o l o g i c a l s e c t i o n s, t h e t i s s u e blocks a r e sectioned t r a n s v e r s e l y s o that, when whole muscle cross s e c t i o n a l a r e a i s multiplied by packing d e n s i t y of myofibers per u n i t area, t h e apparent number i s overestimated by a f a c t o r which i s proportional t o t h e angle a t which myofibers a r e transected i n t h e whole muscle cross s e c t i o n a l a r e a Thus, i f t h i s method i s used it i s a l s o necessary t o measure t h e angle of Inyofiber t r a n s e c t i o n i n whole muscle cross s e c t i o n a l areas and demonstrate t h a t e t cetera it i s constant a t d i f f e r e n t ages, in d i f f e r e n t breeds, This argument i s presented diagramatically i n figure 2 The geometry of t h e problem i s discussed i n d e t a i l elsewhere (Swatland, 1975a) Problem 2 Most researchers who have investigated myofiber numbers i n meat animals have not been unduly concerned with t h e f a c t t h a t t h e a p p r e n t myofiber number (as seen i n muscle cross s e c t i o n s ) m i g h t be l e s s than t h e r e a l number (present i n t h e whole muscle) How many myofibers a r e t h e s e researchers missing? I n t h e case of t h e longissimus muscles of market weight pigs, it has been shown t h a t a MINIMUM estimate f o r t h e number missed is 88% of t h e r e a l number of approximately six and h a l f million qy9f i b e r s (%ratland, 1975b) Problem 3 Problem 2 gives r i s e t o a further question of p a r t i c u l a r importance t o s e l e c t i o n theory Is t h e apparent/real f r a c t i o n fixed or variable? If it is fixed, then carcasses from animals with a n increased r e a l number due t o s e l e c t i v e breeding w i l l have an increased apparent number of myofibers i n t h e i r l o i n chops If it is variable, then an increased r e a l number may or may not cause an increase i n a p p r e n t numbers In a d e t a i l e d study of a small number of animals (Swatland, 1975b) t h e apparent/real f r a c t i o n in t h e a n i m l s examined was not f i x e d In other words, pigs which had t$e most myofibers i n t h e i r pork chops had a high a p p a r e n t / r e a l f r a c t i o n and not a high r e a l number of myofibers This finding does not disprove t h e claims made by Staun and Goldspink, but it does i n d i c a t e t h a t t h e problem i s a l i t t l e more complex than t h e s e researchers may have r e a l i z e d Problem 4 This problem arose from t h e observation t h a t, while m i n i m e s t i mates of r e a l number were not correlated with muscle weight, t h e a p m r e n t number w a s (Swatland, 1975b ) This observation was d i f f i c u l t t o explain on t h e b a s i s of t h e simple t h e o r i e s advocated by Staun and Goldspink However, when t h e angle of t r a n s e c t i o n of myofibers i n whole longissimus muscle cross sections was again taken i n t o account an explanation became evident If f a s c i c u l i continue t o grow i n length a f t e r longitudinal growth of t h e v e r t e b r a l column has slowed down and ceased, then t h e apparent number of nryofibers might a l s o continue t o

6 ANGLE = 45" 7Rfl)NSVE RSE S EC710 N M IC ROTO h

7 increase, as shown i n f i g u r e 3 Strong support f o r t h i s p o s s i b i l i t y was derived by studying l o n g i t u d i n a l f a s c i c u l a r growth Here it was found t h a t p e r s i s t a n t l o n g i t u d i n a l f a s c i c u l a r growth enabled continued growth i n depth of t h e whole muscle cross s e c t i o n, a s shown in f i g u r e 3 Thus, as f a s c i c u l i grew i n length, t h e a p p a r e n t / r e a l probably increased Summary of Problems 1, 2, 3 and 4 ( i ) It is t h e apparent number r a t h e r than t h e r e a l nuiijber of myofibers which is a c t u a l l y present i n t h e eye muscle of a pork chop ( i i ) Staun and Goldspink suggest that w e can increase meat y i e l d by increasing r e a l numbers of myofibers (iii) This m y be possible, b u t, i n t h e porcine longissimus muscle, only i f t h e a p p a r e n t / r e a l f r a c t i o n remains constant ( i v ) An e a s i e r a l t e r n a t i v e method is now evident by which an increase i n t h e eye muscle a r e a of pork chops might be produced and t h a t is t o increase t h e a p p a r e n t / r e a l f r a c t i o n by s e l e c t i n g f o r increased f a s c i c u l a r length Fascicular length i s c l o s e l y r e l a t e d t o eye muscle depth (Swatland, 1976a) and t h i s can be determined by u l t r a s o n i c probing Simple Muscles Arguments have been presented here in an attempt t o show t h a t t o increase t h e appwent numbers of myofibers in pork chops by s e l e c t i v e breeding is not a simple e x e r c i s e B u t w h a t of t h e simpler muscles, w i l l t h e proposed method of Staun and Goldspink work with these? A fundamental point that i s e s s e n t i a l t o t h e i r proposed method, is t h a t r e a l numbers of myofibers must remain constant a f t e r b i r t h Stickland and Goldspink (1973) have described some very small muscles of t h e f o o t i n which t h e apparent number was thought t o be equal t o t h e r e a l number No s i g n i f i c a n t evidence f o r an age o r growth r e l a t e d change w a s detected It is an elementary point, but one t h a t needs t o be emphasized; f a i l u r e t o d e t e c t a change is not a proof t h a t a change does not occur F'urthermore, it should be noted that p o s t n a t a l increases i n r e a l qyofiber numbers have been detected i n c e r t a i n muscles of t h e rat (Rayne and C r a w f ord, 1975 ) The Porcine Sartorius--a "simple" muscle Stickland and Goldspink (1973) e s t i m t e d apparent myofiber numbers i n t h e p o r c h e s a r t o r i u s and detected no change during growth t o an age of 215 days This corresponds, more or l e s s, t o t h e general findings of other researchers who have looked a t s a r t o r i u s muscles in other

8 93

9 94 mamls The sartorius is a good choice for a sample muscle since its fasciculi are almst parallel and, if there are complications due to fascicular arrangement, they are not immediately evident as in the longissimus dorsi The porcine sartorius muscle does, however, contain intrafascicularly terminating my'-(swatland and Cassens, 1972) Being located within the commercially important muscle mass of the butt end of the ham, the sartorius may well behave in the same way during growth as its Larger and more important anatomical neighbors According to the concept of the role of myofiber numbers in muscle growth advocated by Staun and Goldspink, the situation in the sartorius is straight forward; the real number of myofibers increases prenatally, is fixed postnatally and is closely correlated to the real number of myofibers in other muscles This might be a tenuous extrapolation to make from a few observations on apparent qyofiber numbers in postnatal pigs Subsequent studies on this muscle provided grounds for these misgivings Problem 5 In agreement w ith the theories of Staun and Goldspink, the apparent number of myof ibers at the midlength of the porcine sartorius was found to show a dramatic increase during prenatal developmmland, 1973) In disagreement, however, it was also observed, on the basis of very limited data (two animals), that the increase continued postnatally into the neonatal period Later observations based on a larger number of neonates (E animals) confirmed this initial observation (Swatland, 1975~) The theoretical contradiction that this created to the work of Staun and Goldspink was not very important since, whether apparent numbers became fixed at the time of birth or shortly afterwards in the neonatal period, at first appeared to be a technicality It is, unfortunately, a technicality of considerable importance If qyof iber numbers are completely determined prenatally and fixed at the time of birth then it is not unreasonable to assume that, provided the sow is able to sustain all her fetuses with adequate nutrition and in a fairly constant uterine environment, myofiber numbers will be completely determined by the fetal genotype This situation would make selection on the basis of myofiber numbers particularly effective However, what will happen if apparent myofiber numbers are still increasing in the neonatal period? The two most important effects might be (i) a marked increase in the non-genetic component of variation because of the hazardous environment of the neonatal pig with regard to the provision of warmth and food and (ii) an increase in variation of apparent numbers of myofibers due to degree of maturity at birth or early postnatal growth Problem 6 From the work of Staun and Goldspink it can be predicted that apparent numbers of myofibers will remain constant once the neonatal increase is complete Furthermore, it can be predicted that plane of nutrition during growth should have no effect on the apprent rider

10 95 of myofibers at the sartorius midlength However, in a study of 16 Canadian Yorkshires growing at a typical rate to market weight it was found that the apparent number of myofibers at the muscle midlength showed a significant increase (P 005) (Swatland, report in preparation) This initial finding was confirmed in a further study in which 11 animals were placed on a maintenance diet which prevented further growth for 28 day periods prior to slaughter This treatment arrested the increase in apparent number of myofibers predicted from the regression line for full fed animals These findings are difficult to reconcile with the work of Staun and Goldspink Perhaps there are fundamental differences between different breeds of pigs raised in different localities or perhaps there are logical or technical fallacies in the methods used to estimate myofiber numbers Myof iber numbers --conclus ion Apparent numbers of myofibers in the muscles of meat animals may equally well increase (Swatland, 1975~) or decrease (Bendall and Voyle, 1967) during postnatal growth They might also remain constant in certain circumstances Only in these latter circumstances will selection on the basis of myofiber numbers be possible in norm1 pigs Radial myof iber growth Bovine muscle was first examined microscopically in 1714 by Antony van Leeuwenhoek (Liddell, 1960) By the middle of the last century there was still some confusion relating to the extent to which increases in numbers of myofibers and increases in myofiber size accounted for the increase in muscle mass observed during postnatal growth (Quain, 1856) By the end of the century, however, the problem had been more or less solved and it was thought that increases in the bulk of individual qofibers accounted for most of the enlargement of entire muscles observed postnatally (Schafer, 1898) In the early years of the present century Heidenheln (1913) established that myofibrils increased in number by longitudinal division during the growth in size of individual myofibers The extensive and laborious studies of Hammond (1932) provided substantial quantitative confirmation of the changes in myof iber diameters occuring during postnatal growth The literature published in the 40 year perid following Hamond's work in 1932 was reviewed by Hegarty (1971) One of the more recent developments in this area rehtes to the heterogeneity of myofiber growth patterns Before the application of histochemical methods to the study of meat animals by Beecher -- et al (1965) and Moody and Cassens (l9@), myofiber diameters were usually measured as a single population Since unimcdal distributions with regard to diameter were usually found, even researchers who were aware of the physiological differentiation between red and white myofibers (Hmnd, 1932) saw no reason to regard radial growth as being anything other than a uniform process Recent evidence, however, suggests that, in some situations, radial growth of myofibers may not be uniform

11 -- Johnston e t a l (1975) studied t h e diameters of d i f f e r e n t h i s t o chemical types of myofibers i n c a t t l e slaughtered a f t e r 153 o r 233 days on a standard f i n i s h i n g r a t i o n One of t h e most i n t e r e s t i n g findings w a s that BR myofibers were t h e only type t o show any marked r a d i a l growth i n t h e e x t r a 80 days on feed I n porcine muscle, t h e p r o l i f e r a t i o n of myofibers keeps pace, more o r l e s s, with t h e r a d i a l growth of q o f i b e r s (Swatland, ~ )and, i n t h i s s i t u a t i o n too, t h e l a t e r stages of growth a r e characterized only by t h e p e r s i s t a n t growth of red myofibers It i s i n t e r e s t i n g t o note t h a t, i n neonatal porcine muscle, red myofibers with a c i d - s t a b l e ATPase i n i t i a l l y l a g behind t h e o v e r a l l population i n r a d i a l growth (Swatland, 1975d) A t t h e present time, t h e r e f o r e, it i s not unreasonable t o propose, as a working hypot h e s i s, that e a r l y p o s t n a t a l muscle growth is characterized by t h e r a p i d r a d i a l growth of white myofibers and t h e slow growth of red myofibers while l a t e r growth i s characterized only by t h e sustained r a d i a l growth of red myofibers This working hypothesis would explain why t h e o v e r a l l r a d i a l growth of myofibers viewed as a uniform population (mean diameter of a l l t y p e s ) may appear t o slow down a s anlnrals grow older ( I d l a m e e t al, 1973; Chrystall e t a l, 1969) o r reach a maximum before attainment of maximum l i v e weisttmoody e t a - * ) Transformations i n Myofiber Histochemistry During P o s t n a t a l Development With c e r t a i n methods applied i n p a r t i c u l a r ways, t h e histochemical d i f f e r e n t i a t i o n of myofiber types can be detected before b i r t h in meat G, 1972; Ashmore and Addis, 1972; animals (Ommer, 1971; Ashmore Swatland and Cassens, 1973; Ashmore e t al, 1973) b u t is not u n t i l t h e neonatal per id t h a t t h e histochemical d i f f e r e n c e s between myof i b e r types become rully defined (Cassens e t al, 1968; Morita e t al, 1970; Cooper e t al, 1970) Even a t t h i s time, however, i n t h e neonatal porcine s a r t o r i u s, a noticeable increase i n t h e percentage of myof i b e r s with a c i d - s t a b l e ATPase (Swatland, 1975d) i s already occurring This m i g h t be r e l a t e d t o t h e i n i t i a l exposure of p o s t u r a l muscles t o t h e e f f e c t s of g r a v i t y Transformations a l s o occur in t h e l a t e r p o s t n a t a l development of m e a t animals and t h e s e m y pose a s e r i o u s and as y e t unsolved problem f o r researchers studying t h e heterogeneity of r a d i a l myofiber growth I n other words, if myofibers d r i f t from one category i n t o another, it i s d i f f i c u l t t o study t h e growth of a s i n g l e consistent subpopulation of myof ibers I n c a t t l e, Waldmn (1967) observed a decrease i n red myofibers a s animals grew t o 1200 l b l i v e wei&t I n pigs, Cooper e t a l (1970) observed a similar t r a n s i t i o n in t h e 24 weeks f o l l a r i n g b i r t h Ashmore e t a l (1972) observed a s i m i l a r phenomenon i n p i g l e t s, lambs and calves with a t r a n s i t i o n of q o f i b e r s from t h e cdi t o t h e aw category In agreement with t h e s e s t u d i e s, Van Den Hende e t a l (1972) presented q u a n t i t a t i v e evidence of a proportional d e c l i n e i n nurdbers of myofibers with a high content of oxidative enzymes in t h e porcine lonaissimus muscle These authors examined Belgian P i e t r a i n and Landrace pigs but a similar t r a n s i t i o n has been described q u a n t i t a t i v e l y i n Canadian

12 97 Yorkshire pigs where, in accordance with Ashmore's theories, the transition only occurred within myof ibers with alkali-stable, acid- Labile ATPase (Swatland, 1976b) Bearing in mind the variations in histochemical methods, both intended and unintentional, between different research groups these findings are in close agreement British Large White pigs, however, although closely related to Canadian Yorkshires, appear to develop differently Davies (1972) found that the proportion of myofibers with a low alkaline ATPase activity and a high content of oxidative enzymes increased rather than decreased during growth Since this study appears to have been conscientiously undertaken, it cannot be lightly dismissed and it serves as a warning that mscle growth in pigs of different breeds raised in different localities m y exhibit some as yet unsuspected differences Whether or not exercise affects the histochemical transformation of porcine myofibers is a difficult question to answer Campbell -- et al (1971) found that twa weeks of exercise increased the area occupied by myofibers with alkaline ATPase in the parcine longissimus muscle The extent to which this change was due to growth-related or exerciserelated radial growth of myofibers was not clear and so the existance of trans itions in myof iber histochemistry could not be established In an extensive study of endurance and sprint running in miniature pigs, Fitts -- et al (1973) were unable to detect transitions in myofiber histochemistry Thus, at the time of writing, there seems to be no evidence that excessive exercise affects myofiber histochemistry in pigs From an agricultural viewpoint, however, it might be more interesting to determine if almost total inactivity due to close confinement of meat anbmls has any effect on rqyofiber histochemistry Proliferat ion of Myofibrils The phenomenon of myofibrillar proliferation can be studied by either light or electron microscopy but both methods present problems By light microscopy, sampling problems are minimized and data can be correlated with the histochemical analysis of adjacent serial sections but the perimeter boundaries of individual myofibrils are difficult to discern Electron microscopy provides a far better resolution of individual qrof ibrils but quantification is difficult because adjacent myofibrils are now often seen to be incompletely seprated, being joined by "bridges" of myof ibrilhr nrrterial (Lawrie, 1974) By light microscopy, myofibrils can be identified as unstained areas when the glycogen in the sarcoplasm is stained by the periodic acid-schiff reaction The mean packing density of myofibrils per unit area can then be determined in muscle cross sections using an oil inrmersion objective and data can be collected from specific histochemical types of myofibers by reference to serial sections The mean packing density of myofibrils can be multiplied by the mean cross sectional areas of myofibers determined by projection, mapping and planimetry to give an approximate estimate of the mean number of myafibrils per histochemical type of myofiber In neonatal pigs, the mean value is approximately 60 nlyofibrils/ myofiber (all histochemical types combined; Swatland, 1975~) There

13 i s some evidence t h a t rapid p r o l i f e r a t i o n of myofibrils occurs in white myofibers of younger animals while only red myofibers have any sustained growth i n heavier a n i m a l s One possible explanation for t h i s i s t h a t t h e continued growth of heavier anirnals r e q u i r e s g r e a t e r p o s t u r a l muscle s t r e n g t h t o overcome g r a v i t y and it seems l i k e l y t h a t red myofibers a r e primarily concerned with p o s t u r a l muscle function A s far as can be ascertained by subjective examination and from data on t h e mean packing d e n s i t y of myofibrils per u n i t area, t h e r a t i o of sarcoplasm t o myofibrils and t h e s i z e s ( i n cross s e c t i o n ) of myofibrils i s more o r l e s s constant t h r a g h a u t later postnatal development This condition i s consistent v i t h theory t h a t q o f i b r i l s p r o l i f e r a t e by l o n g i t u d i n a l f i s s i m once they grow t o a l a r g e s i z e (Heidenhain, 1913; Van Breeman, 1952; Goldspink, 1970) Longitudinal Muscle Growth Lmgitudinalmyofiber growth occurs by t h e formation of new sarcomeres a t t h e ends of e x i s t i n g myofibrils (Muir, 1961; Bendall and Voyle, 1967) In meat animals, longitudinal muscle growth can be studied a t t h r e e l e v e l s of organization, ( i )whole muscle length, ( i i )f a s c i c u l a r length, and (iii)myofiber length Since muscles are extensible it i s necessary t o standardize muscle length by taking samples a f t e r t h e occurrence of r i g o r mortis with carcasses s e t i n a standard posture Alternatively, sarcomere lengths can be measured and used t o a d j u s t t h e d a t a on longitudinal growth t o a constant sarcomere length Whole muscle length may or m y not be c l o s e l y r e l a t e d t o f a s c i c u l a r length and q o f i b e r length For example, if muscle f a s c i c u l i are arranged a t an angle t o t h e l o n g i t u d i n a l axis of a muscle, l a r g e changes i n f a s c i c u l a r length w i l l only produce small or even n e g l i g i b l e changes in whole muscle length Alternatively, muscle f a s i c u l i m i g h t terminate on a connective t i s s u e framework within t h e b e l l y of t h e muscle s o t h a t whole m s c l e length would be proportional t o f a s c i c u l a r length only i f t h e r a t i o of f a s c i c u l a r length t o whole muscle length was constant If new sarcomeres are added a t t h e same r a t e t o both a whole muscle and t o a f a s c i c u l u s s h o r t e r than t h e whole muscle, t h e r a t i o of f a s c i c u l a r length t o whole muscle length i s not constant When f a s c i c u l a r length i s used as a parameter of l o n g i t u d i n a l growth t h e s i t u a t i o n is not much b e t t e r s i n c e myofibers may terminate i n t r a f a s c i c u l a r l y thus c r e a t i n g t h e problem of constancy of p r o p o r t i o n a l i t y i n length encountered i n t h e previous example Except i n smll muscles where t h e r e a r e no i n t r a f a s c i c u l a r l y terminating q o f ibers, it i s v i r t u a l l y impossible t o measure qyof iber length without d i s s e c t i n g out individual myof i b e r s Another problem is encountered i n t h e study of longitudinal muscle growth when metal w i r e s a r e inserted through t h e muscle (Mackay and Harrop, 1969; Crawford, 1954; Muhl and Grimm, 1974) The t i s s u e r e s t r a i n i n g the t r a n s v e r s e l y placed wires which are used t o d e t e c t l o n g i t u d i n a l growth is probably connective t i s s u e r a t h e r t h a n t h e highly mobile muscle elements Thus it i s no s u r p r i s e t o f i n d that t h i s method demonstrates t h a t i n t e r s tit i a l growth (between wires ) r a t h e r than t e r m i n a l growth ( a t ends of myofibers) accounts f o r almost a l l t h e o v e r a l l growth i n length Thus, t h i s f i n d i n g i s d i f f i c u l t t o reconcile with t h e hypothesis t h a t new sarcomeres a r e added a t t h e ends of myofibers An important question remains unanswered How do i n t r a f a s c i c u l a r l y terminating myof ibers grow?

14 99 The question of muscle length m y have an important bearing on animal conformtion, primarily i n beef animals where muscle conformation is v i s i b l e in t h e l i v e a n i m l Implicit in m n y e a r l i e r s t u d i e s is t h e concept that muscularity i s determined by muscle g i r t h - - i t s e l f a function of apparent number of myofibers, myofiber diameters and nonmuscle elements such a s intramuscular adipose tissue Muscle length might a l s o be very important, p a r t i c u l a r l y i n t h e hindquarter muscle mss There is evidence t h a t increased f a s c i c u l a r length i n the muscle can increase t h e depth of t h e l o i n eye Double-muscling i n C a t t l e In a f a i r l y recent review of t h e l i t e r a t u r e on t h e myology of t h i s phenomenon (Swatland, 1974), t h e r e appeared t o be an almost unanimous agreement among d i f f e r e n t researchers t h a t t h e c h a r a c t e r i s t i c muscle enlargement t y p i f y i n g t h e condition was mostly due t o an increase i n number of myofibers Thus, the overall condition might be described as muscular hypertrophy while i t s cause can a c c u r a t e l y be designated as myofiber hyperplasia As far as can be ascertained a t t h e present time, t h e condition arises from an increase in t h e real numbers of myofibers a t an e a r l y period in p r e n a t a l development (Swatland and Kieffer, 1974) The occurence of double-muscling i n sheep (Naerland, 1940) is a p o s s i b i l i t y t h a t i n v i t e s f u r t h e r i n v e s t i g a t i o n by those i n t e r e s t e d i n meat production i n t h i s s p e c i e s S a t e l l i t e Cells With some cautious reservations, it i s now generally accepted that t h e increase i n number of myofiber n u c l e i which occurs during p o s t n a t a l development is due t o t h e mitosis of s a t e l l i t e c e l l s and t h e fusion of daughter c e l l s h t o t h e myofiber (Pkiuro e t a l, 1970) I n t h e f i v e year period t h a t has elapsed s i n c e t h e s e then new conclusions were reported a t t h e 1971 Reciprocal Meat Conference (Swatland, 1971), t h e r e has not e x a c t l y been a g r e a t rush t o i n v e s t i g a t e t h e involvement of s a t e l l i t e c e l l s i n meat animals Thus, l i k e l o n g i t u d i n a l muscle growth, t h i s appears t o be a neglected a r e a p a r t i c u l a r l y s i n c e t h e r e are nuw reasonable grounds f o r i d e n t i f y i n g s a t e l l i t e c e l l s by l i g h t microscopy ( Ontell, 1974) LITERATURE CITED American Society of Animal Science 1974 Protein synthesis and muscle growth A Symposium J Anim S c i 38:1050 Ashmore, C R and P B Addis 1972 P r e n a t a l development of muscle f i b e r types i n domestic animals Proc Recip Meat Conf 25:2ll

15 100 Ashmore, C R, D W Robinson, P Rattray and L Doerr 1972 Biphasic development of muscle f i b e r s i n t h e f e t a l lamb 37:241 Ashmore, C R, G Tompkins and L Doerr 1972 Postnatal development of muscle f i b e r types i n d o m s t i c animals J Anim S c i 34:37 Ashmore, C R, P B Addis and L Doerr 1973 Development of muscle f i b e r s i n the f e t a l pig J Anim S c i 36:1088 Bardeen, C R 1903 Variations i n t h e i n t e r n a l a r c h i t e c t u r e of t h e m obliquus abdominis externus i n c e r t a i n mammals Anat Anz 23: 241 Beecher, G R, R G, Cassens, 11 G Hoekstra and E J Briskey 1965 Red and white f i b e r content and associated post-mortem properties of seven porcine muscles J Food S c i 30:969 Bendall, J R and C A Voyle 1967 A study of the h i s t o l o g i c a l changes i n t h e growing muscles of beef animals J Food Technol 2:259 B u r l e i g h, I G 1974 On t h e c e l l u l a r regulation of growth and development i n s k e l e t a l muscle B i o l Rev 49:267 Campbell, A M, G Onan, D Thorns, W Weirich, J A W i l l, R G Cassens and E J Briskey 1971 The e f f e c t of exercise on muscle ATPase, Histochem Cassens, R G, C C Cooper, W G Moody and E J Briskey 1968 Histochemical d i f f e r e n t i a t i o n of f i b e r types i n developing porcine muscle J Anim Morphol Physiol 15:135 Chrystall, B B, S E Zobrisky and M E Bailey muscle grovth i n swine Growth 33: Longissimus Cooper, C C, R G Cassens, L L Kastenschmidt and E J Briskey 1970 Histochemical c h a r a c t e r i z a t i o n of muscle d i f f e r e n t i a t i o n Develop B i o l 23 :169 Crawford, G N C 1954 An experimental study of muscle growth i n t h e r a b b i t J Bone J o i n t Surg 361~2% Davies, A S 1972 P o s t n a t a l changes in t h e histochemical f i b r e types of porcine s k e l e t a l muscle J Anat 113:213 F i t t s, R H, F J Nagle and R G Cassens 1973 C h a r a c t e r i s t i c s of s k e l e t a l muscle f i b e r types i n the miniature p i g and the e f f e c t - o f t r a i n i n g Can J Physiol Fbarmacol 51:825 Goldspink, G 1970 Morphological adaptation due t o growth and a c t i v i t y p 521 In E J Briskey, R G Cassens and B B h r s h ( e d i t o r s ) The Physiology and Biochemistry of Muscle a s a Food, 2 University of Wisconsin Press, Madison

16 101 Hammond, J 1932 Growth and t h e development of mutton q u a l i t i e s i n the sheep London and Edinburgh: Oliver and Boyd Hegarty, P V J 1971 h s c l e f i b e r growth and development Recip Meat Conf 24:319 Proc Heidenhain, M 1913 Uber d i e Enstenhung der q u e r g e s t r e i f t e n Muskelsubstanz b e i der F o r e l l e Arch Mikroskop Anat 83:427 Huber, G C 1916 On t h e form and arrangement i n f a s c i c u l i of striated voluntary muscle f i b e r s Anat Rec 1:149 Jiminez, A S, G H Cardinet, J E Smith and M 3 Fedde 1975 Evaluation of an i n d i r e c t method f o r estimating q o f i b e r number i n t r a n s v e r s e sections of s k e l e t a l muscle Am J V e t Res 36:375 Johnson, E R and A W B e a t t i e 1973 Variation in muscle f i b r e diameter among s e c t ions and i n t r a s e c t ions and between c o n t r a l a t e r a l muscles i n seven bovine muscles J Agric S c i, Camb 81:9 Johnston, D M, D F Stewart, W G Moody, J Boling and J D Kemp 1975 E f f e c t of breed and time on feed on t h e s i z e and d i s t r i b u t i o n of beef muse-le f i b e r types J A n i m S c i, 40:613 LaFlamme, L F, A Trenkle and D G Topel, 1973 E f f e c t of c a s t r a t i o n of breed type on growth of t h e longissimus muscle i n male c a t t l e Growth 37: 249 hwrie, R A 1974 Meat Science Press, Oxford Le Gros Clark, W E Press, Oxford Liddell, E G T Press, Oxford f i g 3-4 2nd e d i t i o n 1945 The tissues of t h e bodr 1960 The discovery of r e f l e x e s Pergamon p 113 p 10 Clarendon Clarendon 1970 Regenerat ion of h u r o, A, S A Shafiq and A T Milhorat Proceedings of t h e Muscular S t r i a t e d Muscle, and Myogenesis Dystrophy Conference, New York 1969 Amsterdam:Excerpta Medica Mackay, B and T J Harrop 1969 An experimental study of t h e l o n g i t u d i n a l growth of s k e l e t a l muscle i n t h e r a t Acta Anat 72:38 Miller, L R, V A Garwood and M D Judge 1975 Factors a f f e c t i n g porcine muscle f i b e r type, diameter and number J Anim S c i 41:66 Moody, W G and R G Cassens 1968 Histochemical d i f f e r e n t i a t i o n of red and white muscle fibers J Anirn S c i 27:961 Moody, W G, D A Tichenor, J D Kemp and J D Fox 1970 Effects of weight, c a s t r a t i o n and rate of gain on muscle f i b e r and f a t c e l l diameter in two bovine muscles J Anim S c i 31:676

17 102 Morita, S, C C Cooper, R G Cassens, L L Kastenschmidt and E J Briskey 1970 A h i s t o l o g i c a l study of myoglobin i n developing muscle of t h e p i g J Anim S c i 31:664 Muir, A R 1961 Observations on t h e attachment of myofibrils t o t h e In J D Boyd, F R sarcolemma a t t h e muscle-tendon junction Johnson and J D Lever, Electron Microscopy i n Anatomy Edward Arnold, London Muhl, Z F and A F G r i m m 1974 Longitudinal growth of s t r i a t e d muscle : a radiographic study Growth 38 :389 Naerland, G 1940 Forekommer dobbeltlenderkarakteren hos andre husdyrarter em s t o r f e? Universe11 hy-perplasi av s t a m e n s og lemmens muskulatur hos sau Skand VeTidskr 30:811 Needham, A E 1964 The growth process in animals Sir Isaac Pitman & Sons, Ltd p 84 London, Omer, P A 1971 Histochemical d i f f e r e n t i a t i o n of s k e l e t a l muscle f i b e r s i n t h e bovine f o e t u s Ekperientia 27:173 Ontell, M 1974 Muscle s a t e l l i t e c e l l s : a validated technique f o r light microscopic i d e n t i f i c a t i o n and a q u a n t i t a t i v e study of changes i n t h e i r population following denervation A n a t Rec 178:211 Quain, J 1856 Elements of Anatow, p 159 Vol 1 6 t h e d i t i o n Edited by W Sharpey and G V E l l i s Ondon, Walton and Maberly Rape, J and G N C Crawford 1975 Increase i n f i b r e numbers of t h e r a t pterygoid muscles during p o s t n a t a l growth J Anat ll9:347 Schafer, E A 1898 Fibres: t h e i r f i g u r e and measurement In Quain's Elements of Anatomy Edited by A E Schafer and G D Thane Vol 1 P a r t 11 pp e d i t i o n London:Longmans Schieff erdecker, P 2891 Morphologie des Muskelgewbes : Bau der h s k e l n I n "Die Gewebe des menschlichen Korpers and i h r e mikroskopische Untersuchuny" by W Behrens, A Kossel and P Schiefferdecker Zweiter Band, Gewebelehre Braunschweig: Harald Bruhn Staun, H 1972 The n u t r i t i o n a l and genetic influence on number and s i z e of muscle fibres and t h e i r response t o carcass q u a l i t y i n p i g s World Rev Anim Prod 8/3, 18 Stickland, N C and G Goldspink 1973 A possible i n d i c a t o r muscle f o r t h e f i b r e content and growth c h a r a c t e r i s t i c s of porcine muscle h i m Prod 16:135 Swatland, H J 1971 Morpholoa of t h e moror innervation of muscle Proc Recip Meat Conf 24:400

18 Swatland, 3 J 1973 Muscle growth i n t h e f e t a l and neonatal p i g J Anim S c i 37:536 Swatland, H J 1974 Developmental d i s o r d e r s of s k e l e t a l muscle in c a t t l e, pigs and sheep Vet Bull 44:179 Swatland, H J 1975a Morphology and development of endomysia1 connective t i s s u e i n porcine and bovine muscle J h i m S c i 41: 78 Swatland, H J 1975b Effect of f a s c i c u l a r arrangement on t h e apparent number of qyofibers i n t h e porcine longissimus muscle J h i m S c i 41:794 Swatland, H J 1975~ Myof i b r e number and myofibrillar development i n neonatal p i g s Z b l Vet bled A 22:756 Swatland, H J 1975d Histochemical development of myofibers i n neonatal p i g l e t s Res Vet S c i 18:253 Swatland, H J 1976a Longitudinal f a s c i c u l a r growth i n t h e porcine longissimus muscle J h i m S c i 42:63 Swatland, H J 1976b p i g s Z b l Vet Med changes during muscle growth AHistochemical In press in Swatland, H J 1976~P r o l i f e r a t i o n of myofibrils i n t h e porcine longissimus muscle J Anim S c i In p r e s s Swatland, H J and R G Cassens 1972 Muscle growth: t h e problem of f i b r e s with an i n t r a f a s c i c u l a r termination J Anim S c i 35:336 Swatland, H J and R G Cassens 1973 P r e n a t a l development, h i s t o chemistry and innervation of porcine muscles J Anim S e i 36:343 Swatland, H J and N M, Kieffer 1974 F e t a l development of t h e S c i 38:752 double musc1ed condition i n c a t t l e J:Anim Van Breemn, V L 1952 Myofibril development observed with t h e e l e c t r o n microscdpe Anat Rec 113:179, E Muylle, W Oyaert and P De Roose 1972 Changes i n muscle c h a r a c t e r i s t i c s in growing pigs Histochemical and e l e c t r o n microscopic study Zbl Vet Med A l 9 : l O Z Van Den Hende, C Waldman, R C 1967 Changes i n red f i b e r content of bovine muscle with increasing age Special Report, University of Wisconsin, Madison, Cited by R G Cassens, C C Cooper and S Morita 1969 D i f f e r e n t i a t i o n of muscle f i b e r s during growth and development Proc Recip Meat Conf 22, p 117 c

19 104 Dave Anderson: Thank you, Howard, for that stimulating discussion on Postnatal Muscle Development If you have any questions for Howard or any of the other speakers, please hold them until the discussion period at the end of today's session We are privileged to have with us as our second speaker, Dr Richard Keesey, Professor of Psychology, University of Wisconsin Dr Keesey received his PhD from Brown University in 1960, took a Post- Doctorate at the Brain Research Institute, UCIA Since that time, he has been at the University of Wisconsin During the last five to ten years, Dr Keesey and his students have studied hypothalamic regulation of food intake and body weight We are privileged to have h3m share with us today some of the results of that work Dr Keesey