EFFECT 0? THE DM’S WEIGHT GATN DURING GESTATTON 0N LITTER 812E ANO THE 011 LIZATTON 0* WAme F01? THE COLLECT10N OF UTERiNE FLU1DSTN MNE Thesis $05 the 093m of: M, 8 WWW STATE U’NEVERSETY CLIFFORD E. MGRCDM 111 19175 ,1 f ‘ H I ’ i 4. I . E - f ~ ) 9 s 1 V 1 . .M--§ I 55815 f} 1 B“ K BWBERY “‘18.; L: TRAR’ BINDERS 3;;;;;::nnr Inn-m“ ABSTRACT EFFECT OF THE DAM'S WEIGHT GAIN DURING GESTATION ON LITTER SIZE AND THE UTILIZATION OF LAPAROSCOPY FOR THE COLLECTION OF UTERINE FLUIDS IN SWINE BY Clifford B. Morcom III Two experiments were conducted using sows from the Michigan State University swine barn. The first experiment was conducted to determine if differences in body weight at breeding, body weight gains during each trimester of gesta- tion, and total weight gain during gestation were related to litter size at birth. There was no significant correla- tion between any of these parameters and litter size. A slight trend (p<<0.l7) was found for gain during the first trimester and litter size. The second experiment dealt with weight gain during the first twenty-five days of gestation and peripheral plasma progesterone levels. Body weight gain during the initial trimester of gestation appears to peak at day 15 then decrease before rising again. This parallels the peripheral plasma progesterone levels which peak at day 12 then decrease. Clifford B. Morcom III A third experiment led to the development of a tech- nique utilizing the laparoscope for the collection of uterine fluid. Successful collection was accomplished 78% of the time and 40% of the saline flushed into the uterus was recovered. This procedure allows the routine collec- tion of uterine fluids without the trauma and stress of laparotomy. EFFECT OF THE DAM'S WEIGHT GAIN DURING GESTATION ON LITTER SIZE AND THE UTILIZATION OF LAPAROSCOPY FOR THE COLLECTION OF UTERINE FLUIDS IN SWINE BY Clifford B. Morcom III A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Animal Husbandry 1975 ACKNOWLEDGEMENTS Sincere thanks is given to Dr. W. R. Dukelow for his help and patience in guiding me to the completion of this work. Also to the entire Endocrine Research Unit and in particular David A. Snyder and Dr. David E. Wildt. Finally, appreciation is expressed to Dr. Ellwyn Miller and the Michigan State University Swine Barn for the use of their animals. ii TABLE OF INTRODUCTION.......... REVIEW OF LITERATURE........ weight Gain000000000... Laparoscopic Technique. MATERIALS AND METHODS....... Experiment 10.0.0000... Experiment 2......... Experiment 3......... RESULTS...0.0000.0000 ........ 0.0.. weight Gain.00000 ....... .0000 Uterine Fluid........ DISCUSSION0000.000.000.00. SUMMARY AND CONCLUSION.... LITERATURE CITED.......... CONTENTS APPENDIX00000000000000000 00000000000000000 .0... VITA...0.0.0000 00000000000000 0...... ......... 0000... iii Page 21 21 22 23 28 28 41 43 49 51 6O 62 TABLE II. III. IV. VI. LIST OF TABLES Maximum, Minimum and Average Weights for 46 Sows on Experiment 10...... ...... 0.00.00.00.000000000 Litter Size, Breeding Weight, Weight Gain During the First 30 Days of Gestation, and Total Weight Gain During Gestation, Listed According to Litter Size and Parity.......................... Average Weight Gain of Sows and Litter Size Listed According to Breed and Parity............ Body Weights and Percentages of Day 3 Weight of Sows Injected with Progesterone-estrone for 10 Days Beginning Day 13 and Ending on Day 22 of Gestation....................................... Body Weights and Percentage of Day 3 Weight of Control Sows Bled............................... Actual Weights and Percentage of Day 3 Weight of Control Animals Not Bled........................ iv Page 29 30 34 35 36 37 LIST OF FIGURES FIGURE Page 1. The upper left part of the picture shows the grasping of the uterus with laparoscopic forceps and the insertion of a 15 ga. needle through which the polyethylene tubing is fed. The lower right corner shows the tubing in place. The circle indicates the end of the tubing which is blocked and small perfora- tions along the side... ..... .................. 25 2. Average weight gain of sows during gestation. (N: 46). ooooooooooooooooooooooooooooooooo o... 32 3. Control sows bled. (N = 6) .................... 38 4. Sows injected with progesterone/estrone. (N=6) 000000000 o 000000000 coco. ..... a... ...... 39 5. Control sows weighed during first 25 days of gestation. (N = 7) ........................... 40 INTRODUCTION The heritability of litter size and other reproductive traits is low, indicating that environmental factors have a large bearing on litter size and reproductive performance. It seems reasonable that the uterine environmental condi- tions which a sow creates for her litter could influence the litter size. It is also possible that the environmental conditions that the sow creates for the developing fetus may reflect her overall condition and weight changes during gestation. One purpose of the present experiments was to determine if litter size was affected by 1) initial weight at breeding, 2) rate of gain during each trimester, and 3) total gain during gestation. It has been reported by Reddy 35 a1. (1958) and Wildt (1975) that a progesterone-estrone combination, when admin- istered at certain times during gestation, causes an increase in litter size. Since trends have been reported indicating that increases in the dam's weight during the first few weeks of gestation lead to increased litter size, we tested to see if the progesterone-estrone injections affected the dam's weight changes during this period. In studying the phenomenon of embryonic mortality, particular emphasis has been placed in the initial stages of gestation when the greatest proportion of the mortality occurs. The uterine environment, and the different proteins associated with it, are implicated in this problem. Thus, there is a need for a simple procedure for regularly col- lecting uterine fluids from the pig, with minimal stress. This was accomplished using the laparoscope. REVIEW OF LITERATURE The following literature review will deal in two major areas: 1) Body weight gain of a sow or gilt during gesta- tion, and 2) Laparoscopy and its utilization among various research animals. Furthermore, the body weight gain will be broken down into six subgroups: 1) body composition of the pregnant animal; 2) general effects of weight gain dur- ing pregnancy; 3) body weight and its effects on puberty; 4) the effect of varying feed intake; 5) dietary protein, and 6) amount of dietary energy on reproductive performance. Weight Gain 1) Body Composition Weight gains of the pregnant sow or gilt may be broken down into three parts: l) gain of the fetus, uterus, and mammary tissue; 2) normal body weight gain of an animal fed in excess of the requirements for the non-pregnant sow; 3) and pregnancy anabolism, which is body weight gain of the pregnant animal given feed only sufficient for mainte- nance of a non-pregnant animal. This means that a ration that just maintains an animal when she is not pregnant, if fed to her while she is pregnant, will allow her to gain weight. The reason for the latter is unclear, but may be due to an increase in the efficiency of energy utilization (Lodge gt gt., 1966b). Heap and Lodge (1967) compared the body composition of pregnant and non-pregnant sows fed equal quantities of the same diet. The pregnant sows gained an average of 35.9 kg, while the non—pregnant sows gained only 4.3 kg. The extra weight of the pregnant animals was accounted for almost entirely by reproductive organs and extra carcass weight. The extra carcass weight was from muscle, fat and mammary tissue, with the extra muscle and fat being mainly in the abdomen. However, the percent of fat was significantly greater in the back and hams of the pregnant group. 2) General Weight Gain Body weight gain of the dam during pregnancy, and its effect on litter size has been studied by McKenzie (1928), Zeller gt gt. (1937), Stewart (1945), Bowman gt_gl, (1961), Bowland (1964), Lodge gt gt. (1966b), and Buitrago gt gt. (1974). McKenzie (1928) presented data showing an increase in litter size from sows with a larger daily weight gain for the first four weeks of gestation. A tendency towards larger litters with increased rate of weight gain of the dam for the entire gestation was shown by Zeller (1937), Stewart (1945) and Bowland (1964). Bowland's experiment utilized four groups of gilts fed differently during growth and gestation. The groups were analyzed for within group and between group correlations of gestation gain and litter size. The gilts within each group which gained the most during gestation had the largest litter size, the correla- tion being 0.24. However, when comparing the different groups, the groups which had gained the least had larger litters. The correlation between gestation gain and litter size was larger and negative (-.74) than the within group correlation (0.24). From these results he feels, within limits, that feeding methods should be employed which pro— duce low gestation weight gains. The weight of the sow or gilt at breeding affects ovulation rate and thus has the potential to affect litter size. Positive correlations between ovulation rate and the weight of the gilt at breeding were shown by Gossett and Sorensen (1959a) and Young gt_gl, (1974). It should be noted, however, that litter size, measured at day 40 of gestation, was not correlated with the body weight of the gilt at breeding (Gossett and Sorensen, 1959a). With sows, increased body weight at breeding raised ovulation rate and increased litter size (Heap gt gt., 1967). Bowman gt gt. (1961) found that steadily increasing the mean breeding weight of the sow to a maximum of 471 1b. caused an increase in litter size. The maximum number of pigs weaned were from sows weighing 354 lb. at breeding. If the trends reported above hold true, will a fat sow have a higher reproductive performance than a leaner sow? Pomeroy (1960) notes that excessive fatness is often associ- ated with reproductive failure by commercial pig producers. The sterility is not caused by obstruction of the oviducts. Many of these overly fat animals fail to show estrus, and he suggests that estrogen may be absorbed by the fat which prevents estrous behavior. Also, Caldwell gt gt. (1972) states a tendency for fatter sows to farrow fewer pigs. At the other extreme, Pike and Boaz (1972) had one group of sows with low fat reserves which gave fewer fetuses (10) than sows on other treatments (13.5) and they cite a prediction by Elsley gt gt. (1968) that reduced sub-cutaneous fat may reduce reproductive performance. Lodge (1969) found a tendency for sows on a high dietary allowance during pregnancy to reach earlier and lower peaks in litter size than animals fed lower allowances during pregnancy. Thus a sow maintained on a lower dietary ration will produce smaller litters in her first few parities, but larger litters in subsequent parities, than a sow maintained on a larger dietary ration. 3) Attainment of Puberty Body weight is also correlated with the gilt's age at puberty. A negative correlation exists between age and weight at puberty (Robertson gt gt., 1951; Squiers gt gt., 1952). Warnick gt gt. (1951) also found this negative correlation between age and weight at puberty within breed- ing lines. However, heavier breeding lines (body weight measured at 154 days of age) tended to reach puberty slower. He feels genes which bring about growth tend to slow sexual development, but disease and other environmental factors can mask this correlation. The amount of nutrition the growing animal receives affects the age the animal attains puberty. Zimmerman gt gt. (1960), Self gt gt. (1955), Gossett and Sorensen (1956,1959a) found that gilts receiving more feed while growing attain puberty later than those fed a lighter ration. Self gt gt. (1955) concluded that the fatness of the animal, as condi- tioned by the level of feeding, may affect the attainment of puberty. In contrast, Haines gt gt. (1959) and Zimmerman gt gt. (1960) found that gilts receiving greater nutrition during growth attained puberty earlier. However, in compar- ing the weight at puberty of the gilts Haines had on full feed, with gilts Self gt gt. (1955) had on the full feed, Haines‘ group weighed considerably less, much closer to what Self's two-thirds full feed group weighed at puberty. This indicates they never achieved the degree of fatness Self's full-fed group did. Finally, Sorensen gt gt. (1961) and Robertson gt gt. (1951) found the amount of feed during growth did not affect age at puberty. However, Robertson reported that the amount of protein did affect the age at puberty with a high protein diet increasing the age of the animal at puberty. 4) Effects of Varying Feed Intake One direct method of affecting the weight of a sow or gilt during gestation is to vary the level of feed. Lodge gt gt. (1966) fed three groups a 14.1% protein diet at the following levels: Group A, 6 lbs per day; Group B, 3 lbs. per day; and C, 3 lbs. per day until day 76 of gestation when it was followed by 6 lbs. per day until farrowing. There was a significant difference between the average net gain of each group. The treatments did not influence pre- natal and post-natal mortality; however, there was a sig- nificant affect on birth weight with lighter pigs born to sows receiving less feed. This agrees with the findings of O'Grady in 1967, Baker $2.31: (1969) and Hesby gt gt. (1972). In 1969, Baker gt gt. showed that the lowest amount of feed (0.9 kg/pig/day) had a significantly lower farrowing per- centage than the higher levels, but there was no difference in litter size. The weight gain during gestation increased quadratically as the gestation diet increased from 0.9 kg/pig/day to 3.0 kg/pig/day. Self gt gt. (1960) found a consistent failure of their full-fed group to perform as well as the average of their two limited—fed groups (two—thirds full—feed and one-third full-feed) in farrowing over a 3 year period. This indi- cates that larger, heavier gilts and sows produced on a higher level of feed have a lower reproductive performance. This agrees with Self gt gt. (1955), Gossett and Sorensen (1959), Haines gt gt. (1959), and Sorensen gt gt, (1961). Omtvedt (1966), feeding full—feed and restricted amounts of the same diet to both sows and gilts found that gilts far- rowed larger litters when fed the restricted diet. With the sows he found no difference in litter size between the two levels of feed. He notes that sows and gilts on re- stricted diets tend to have slightly more pigs at 21 days and at weaning. The 47 lb. difference in body weight be- tween sows on the high compared to the low feed intake may have been a contributing factor. Waldorf gt gt. (1958), with limited (two-thirds full- feed) and full-fed gilts, showed an absence of a feed effect on fetus weight at 25, 70, and 105 days, even though the feeding regimen did affect the dam as shown by differences in carcass length, weight, and back-fat thickness. This indicates the litter is able to compete with the dam for nutrients, and the dam will suffer rather than the fetus in cases of a moderate shortage of nutrients. Litter size shows a trend toward direct positive effects on fetus weight at day 25, but the number of pigs per uterine horn appears to have a direct negative effect on individual fetus weight as gestation advances. 10 Lodge gt gt. (1966), using sows, and Bowland (1964), Meade gt_gt. (1966), Vermedahl gt gt. (1969), and Buitrago gt_gt, (1974), using gilts, found that animals which gained least during gestation tended to lose less weight at farrow- ing and during a 3-week lactation period. Lodge gt gt. (1966) found this even though his sows consumed about the same amount of feed during lactation. Salmon-Legagneur and Rerat (1962) reported the tissue laid down during pregnancy is labile and is the first to disappear during lactation. Baker gt gt. (1970b) states "weight gain during gestation is an important determinant of weight changes of both the dam and her litter during lactation." Smith (1960) limited the body weight gain of the dam during pregnancy by controlling the intake. For example, 2nd litter sows listed in "good condition" were fed to gain 130 lbs. during gestation while those in "low condition" were fed to gain only 60 lbs. Sows in low condition at farrowing lost less weight and produced less milk with a lower energy content than sows in good condition. In earlier work, Smith (1959) showed a reduction in milk yield by sows fed less during lactation but this was offset by increased energy content of the milk. The percent fat in the milk increased and the lactose content in the milk decreased as "the amount consumed during lactation was lowered. Sows fed Lless during lactation used their feed energy equivalent more Eifficiently than those on a higher plane of nutrition. ll 5) Protein Variations in the Diet Protein is another variable in the diet which can affect weight gain during gestation-—especially in the gilt (Rippel gt gt., 1956). Holden gt gt. (1968) and Hesby gt gt. (1972) reported sow weight gains during gestation to in- crease significantly with increased protein in the diet. They all found that litter size, live pigs farrowed, and birth weight of live pigs was not influenced by different levels of protein. This is in agreement with Clawson gt gt. (1963), Pond gt gt. (1968) and Hawton and Meade (1969). Holden gt gt. (1967) maintained sows on 8, 12, 16, and 20% protein diets through four reproductive cycles. There were no significant differences in the number of live pigs far- rowed, birth weight, or number weaned at 2 weeks. Pond's treatment was the most severe as they reported normal repro- duction from gilts fed a protein-free diet from day 24 of gestation to parturition. This experiment indicates that even severe restrictions of protein will not effect the fetus; therefore, it must compete very favorably with the dam for amino acids even to the point where she will sacri— fice much of her own tissues to support the fetus. Sows on a protein—free diet through most of gestation had lighter pigs. Of the three gilts on a protein-free diet for the entire gestation, two died of peptic ulcers and the other farrowed nine pigs. Jones and Maxwell (1974), feeding three 12 different levels of protein to gilts, linearly increased the number of corpora lutea with increasing protein. However, embryo survival as measured 30 days after breeding was not affected. There has been some contradictory evidence concerning the effect of protein. Baker gt gt. (1974) found an in- crease in litter size from gilts and sows when he varied the diets during gestation with the critical difference being a 16% protein diet compared with a 12% protein diet over the last trimester of gestation. Hawton and Meade (1971) found a trend in the number of pigs farrowed per litter by gilts in the first reproductive cycle to decrease as the quantity and quality of the protein intake during gestation was lowered—-the total difference being nearly one pig per litter (8.8 vs. 7.9 pigs per litter). This trend was not reproduced with second litter sows. The weight gain of the dams was not affected by treatments during the first reproductive cycle, but was affected during the second ges— tation. Sows receiving high protein (341 g/da) gained sig- nificantly more than sows on low protain diets (154 g/da). Frobish gt_gt, (1966) found that more pigs farrowed on a high rather than low protein intake, but no significant effects were observed on birth weight, gain from birth to weaning, or sow weight changes from the start of the trial to weaning. 13 Another effect of low dietary protein may be an increase in the number of anestrous gilts. Jones and Maxwell (1974) reported a trend toward an increased number of anestrous gilts with a lower level of protein in the diet. The per- centage of anestrous gilts not exhibiting a third estrous cycle for the three groups fed 143, 254, and 345 g. of pro- tein was 20.7, 23.3, and 3.6% respectively. Similar trends were reported by Adams gt gt. (1960), Boaz (1962) and Pond gt gt. (1968). Low protein diets during gestation exert an effect dur- ing lactation in spite of adequate diet for the sow during lactation. Hesby gt gt. (1972) found the weight of baby pigs from birth to weaning tended to increase as dietary protein to the sow during gestation increased. This is in agreement with Rippel gt gt. (1965) and Holden gt gt. (1968). Stevenson and Ellis (1957) showed by using 20 or 17.5% crude protein rather than a 15% diet during gestation the viabil- ity and survival rate of the baby pigs increased. Frobish gt gt. (1966) also showed a significantly greater number of pigs weaned by sows fed a high protein rather than a low protein diet during gestation. Baker gt gt. (1970a) showed sows weaned significantly more pigs when fed a corn-soybean (12, 16, and 20% crude protein) or an opaque —2 corn (9.7% crude protein) than when fed corn (8.8% crude protein) dur— ing gestation, showing the inability of gilts fed a corn l4 diet during gestation to perform at an optimal level during lactation. The increased litter performance is probably due to the better milking ability of a sow or gilt on a high protein diet. It has been suggested that the "protein anabolism" in sows during pregnancy is available as labile protein for lactation purposes (Pond, 1973). Holden gt gt. (1967 and 1968) found a significant rise in milk protein with increased protein in the diet. Strothers and Milne (1964) fed diets during gestation containing 8 or 14% pro- tein and a 14% protein diet to all sows during lactation. At the end of the first week, the milk from the sows on the high level of protein during gestation, contained more crude protein than milk from sows fed the lower level during gestation, but this effect was not observed at the third week. This indicates a high quality feed after parturition cannot make up entirely for a poorer diet before parturi- tion. Baker gt gt. (1970) reports gilts on a higher protein diet either for the entire gestation or just the last tri- mester lost more weight during lactation than gilts on a lower protein intake, indicating heavier milking. Hawton and Meade (1971) noted body weight loss of dams during lactation tended to decrease as the level and quality of the protein during gestation decreased. Pigs from dams fed 250 g/da. of protein during gestation weighed signifi- cantly less at 21 days than those produced by sows fed 341 15 g/da. of protein. Hesby gt gt. (1972) found the percentage of live pigs weaned tended to increase as quality and quan- tity of protein fed during gestation increased. However, this does not mean overly fat gilts at parturition will milk best. Self gt gt. (1960) found the 3—year average weaning weights for total litters was in favor of his limited—fed groups, indicating lactating ability is not entirely depend- ent upon the weight of the dam at farrowing and too much weight may be detrimental. The ability of the dam to recover effectively during the last trimester from a poorer diet during the first two- thirds of pregnancy was shown by Baker gt gt. (1970b). Gilts fed a 16% protein diet following an 8.7% protein diet gained as much weight during gestation as those fed a 12% diet throughout gestation even though they consumed 10% less protein. 6) Amount of Energy in the Diet Energy is another variable in the diet which will af— fect weight gain, ovulation rate, and litter weight. One phenomenon of increased energy is flushing which causes a significant increase in ovulation. The ovulatory response to flushing depends upon the length of time and the point at which it occurs in the estrous cycle. Zimmerman gt gt. (1957 and 1960), Lodge and Hardy (1968), Naber and Zimmerman (1971) and Brooks (1972). 16 According to Daily gt gt. (1972), flushing causes an increase in follicular fluid weight and the number of large follicles if done over the last 4 to 6 days of the estrous cycle. When performed from day 10 to day 15 of the cycle, the weight of the fluid was increased and more medium sized follicles, compared with the number of small follicles were formed, but these effects were lost by the end of estrus. Zimmerman gt gt. (1957) found such individual characteris- tics of gilts as pre—flush and post-flush weight and back— fat thickness, as well as average daily gain and feed con- sumption during the flushing period were not significantly associated with ovulation rate. The usefulness of flushing may be questioned as flush- ing also increases embryonic mortality, and there is no sig- nificant difference between number of embryos found in flushed and unflushed animals (Haines, 1955; Sorensen gt gt., 1961; and Gibson gt_gt., 1963). In a conflicting report Lodge and Hardy (1968) found a significant increase in litter size by flushing with the first feed following breeding. It is obvious that the potential for increased litter size occurs with flushing--the problem is to decrease (embryonic mortality so that the potential is realized. 13azer EE.§£° (1968) found a reduced survival rate of embryos tlransferred from gilts raised on a low level of feed (1.81 l7 kg/pig/da.) to gilts on the same level which had been flushed for 14 days prior to breeding. The data from two other experiments he conducted indicates the embryos are equally viable in flushed and non-flushed gilts, which indi- cates some factor in the uterine environment causing the increased mortality. The amount of energy in the diet throughout gestation was studied by Frobish gt gt. (1966). The two levels of energy in the diet used were 10,800 and 5,400 kcal./pig/day. Sows receiving the low energy diet tended to farrow more live pigs than sows on the high energy diet. By the end of three reproductive cycles, over half the sows on the high energy intake were removed from the experiment because of death or failure to farrow. Of those failing to farrow, large amounts of fatty tissue were found around the repro- ductive organs indicating excessive internal fat. Frobish gt gt. (1973) fed 3,000, 4,500, 6,000, and 7,500 kcal. per day diets through three reproductive cycles and found the total pigs farrowed decreased in a linear fashion with in- creasing energy. The highest energy group (7,500 kcal.) had the fewest number of sows complete three reproductive cycles, with leg abnormalities being the major cause of re— moval. There were significant increases in weight with increases in energy. 18 Gossett and Sorensen (1959b) found low dietary energy for gilts more desirable than high energy diets as a greater number of normal 40 day embryos were present. Gilts fed the high energy diet had a greater embryonic mortality after day 25 than gilts fed the low energy diet. In contrast Frobish (1970) and Vermedahl gt gt. (1969) had increases in the total number of pigs born for gilts on a high energy ration compared with gilts on a low energy ration. The difference between the results in these experi- ments and those conducted by Frobish gt gt. (1966 and 1973) may just indicate the difference in nutritional requirements between gilts and sows. Baker (1971) felt severe calorie restriction causes interference with implantation and reduces the gilt‘s ability to maintain pregnancy. Frobish (1973) lends support to this theory when he found the major cause of removal of sows from his low energy diet (3,000 kcal./pig/da.) was failure to conceive. However, in still another experiment by Frobish (1970) no difference in viable embryos was found at day 30 of gestation between two groups of gilts fed diets containing 6,000 kcal/pig/da. or 2,600 kcal./pig/day. He gives no data on the number of animals which failed to conceive on the two treatments. Several inconclusive studies concerning the amount of energy in the diet and its effect on litter size have been done. Sorensen gt gt. (1961) found no significant differ- ence between two self fed groups of gilts receiving either 19 53,000 or 93,000 kcal./100 lbs. feed, but the trend favored gilts receiving the low energy diet. Clawson gt gt. (1963) had gilts who farrowed significantly more pigs on a low energy diet in one trial, but in the second trial the gilts receiving the high energy diet farrowed more pigs. Finally, Buitrago gt gt. (1974) reported no significant difference between litter size from gilts receiving different amounts of energy in their diets during gestation, but the dams with the lower energy intake during pregnancy tended to have smaller litters. Laparoscopic Technique Utilization of laparoscopy for ovarian examination was developed in human medicine (Semm, 1969; Balin, Wan and Rayan, 1963), but has also been adapted for laboratory and domestic animals. A list of animals on which laparoscopy has been used includes rabbits (Fujimoto gt gt., 1974), goats (Dukelow gt gt., 1971), primates (Dukelow gt gt., 1971; Jewett and Dukelow, 1972; Rauson and Dukelow, 1973; Graham gt gt., 1973; Harrison gt gt., 1974), sheep (Phillippo gt gt., 1971; Seeger, 1973; Snyder and Dukelow, 1974), and swine (Wildt gt_gt., 1973). Laparoscopy allows direct uterine and ovarian observa- tion with a minimal amount of surgical stress and trauma. With some ingenuity and minor modifications or additions in equipment, most of the things now being done by laparotomy 20 could be done by laparoscopy. Murray gt gt. (1972), Chen gt gt. (1973), Knight gt gt. (1974), and Irrtani gt gt. (1974) have all utilized laparo- tomy at some point in this procedure for the collection of uterine fluids. To date there has been no report of a method for the collection of uterine fluid without the trauma of laparotomy. Using the technique of laparoscopy in swine described by Wildt gt al. (1973), a method of uterine fluid collection has been developed. MATERIALS AND METHODS Three experiments were conducted using different groups of gilts or sows. The first experiment dealt with initial body weight and body weight gains during different periods of gestation. The second experiment was concerned with changes in weight and its correlation with progesterone levels during the first 25 days of gestation. The final experiment was concerned with the development of a technique for the routine collection of uterine fluid utilizing the laparoscope. Experiment 1 A total of 107 sows were weighed at two week intervals throughout gestation. The range in parities was from two to eight. Three types of pigs were used, Yorkshire, Hamp- shire, and Yorkshire-Hampshire crosses. Boars of proven fertility were used and breeding was natural except for six sows which were bred artificially using frozen semen. Of the 107 sows, twenty-nine were removed due to an outbreak of transmissible gastroenteritis during the second week in March, 1974. Sixteen were removed due to a faulty scale giving improper readings, six returned to heat, six were sold or removed because of lameness, and four aborted. 21 22 All sows were kept in an environmentally controlled gestation barn at a temperature of 13°C. They were indi- vidually tied or placed in stalls throughout gestation. Feeding took place once daily in the morning, and the sows were fed individually 4 lb of a 13% protein, corn-soybean meal feed fortified with minerals and vitamins. Approxi— mately one week prior to farrowing, they were moved to the farrowing house and placed in individual stalls. While in the farrowing house, sows were fed twice daily 4 lb of a 16% protein, corn—soybean meal ration fortified with Vitamin E and selenium. All weighing was done on two scales located in the farrowing and gestation barns. Experiment 2 Twelve sows were weighed at regular intervals through- out the first twenty-five days of gestation. They were bled five days before breeding and l, 4, 8, l4, 16, 18, 20, 22, 24, and 29 days after mating. Six of these sows were given injections of progesterone and estrone at a ratio of 2000:l (25 mg. of progesterone and 12.5 ug of estrone) on days 13 to 22 of gestation. Day 0 was the day of breeding. The sows were bled from the anterior vena cava using a 16 ga. 11.5 cm. needle. Progesterone assays were carried out on the serum from these sows. The assays utilized a double antibody and were carried out by Dr. David Guthrie at the USDA in Beltsville, Md. Seven additional sows were weighed 23 at regular intervals throughout the first twenty-four days of gestation. Experiment 3 The laparoscopic procedure makes use of a 135° pedia- tric laparoscope 5 mm. in diameter and 24.5 cm. long. Light is transmitted from a Model 4000 projector light source (Wolf Co.) through a flexible fiber—optic cable. The laparoscope is inserted through a 6 mm. cannula equipped with an attachment for abdominal insufflation. A 3 mm. tactile probe is inserted laterally to the laparoscope to manipulate the internal organs. The collection of uterine fluid utilizing the laparo— scope was accomplished with a 15 ga 7.6 cm. needle, P. E. 90 tubing (Clay-Adams), laparoscopic forceps, and a 10 m1. syringe. The technique of laparoscopy in swine has been reported previously from this laboratory (Wildt gt gt., 1973). The gilts were given a tranquilizer, phencyclidine HCl (2 mg./ kg.) followed by the intravenous injection of sodium pento- barbital (5.5 mg./kg.). The use of the tranquilizer phen- cyclidine HCl is a minor revision of the procedure previously reported. It is administered intramuscularly and immobilizes the animal within five to ten minutes. This facilitates the administration of the sodium pentobarbital and allows the lowering of the dosage by one-third (15 mg./kg. to 5.5 24 mg./kg.). Salivation occurs due to the use of the phencycli- dine HCl, but this rarely presents a problem as the slope of the table facilitates drainage from, rather than into, the lungs. However, in animals where congestion leads to difficulty in breathing, 1 mg. of atropine sulfate is ad- ministered intramuscularly. Two incisions, one midventral and the other dorso- lateral, were made and the laparoscope and probe were in- serted. Once the tract was exposed and the tubo-uterine junction was found, the probe was removed and an accessory trocar cannula inserted in the same incision. The trocar was removed and the laparoscopic forceps inserted into the abdominal cavity. The uterus was grasped with the forceps and held in position while a 7.6 cm. 15 ga. needle was in- serted through the abdominal wall at a site above the point of cannulation (Figure 1). After insertion of the needle into the uterine horn, polyethylene tubing, with the termin- al end sealed with small perforations along four centi- meters of its length was inserted through the shaft of the needle into the uterine lumen. Observation of the tubing entering the lumen was possible with the laparoscope by the bulging of the uterus due to the pressure from the tubing. Approximately 7 cm. of tubing was fed into the uterus. The table was lowered to the horizontal position from the 30° angle used in laparoscopy, then 6 m1. of sterile saline was 25 WW ; .............. \\ \ \~ ( l "f, x \‘ . N1 (w Figure l. The upper left part of the picture shows the grasping of the uterus with laparoscopic forceps and the insertion of a 15 ga. needle through which the polyethylene tubing is fed. The lower right corner shows the tubing in place. The circle indicates the end of the tubing which is blocked and small perforations along the side. 26 flushed into the uterus and aspirated. The gilt was re- turned to the 30° angle, and the tract and tubing were observed through the laparoscope to make sure that the tubing had remained in place. The tubing and laparoscope were then removed and the gilt was placed in a heated barn to recover. Furacin ointment and powder were placed on the external incision, and 700,000/units procaine penicillin was injected as prophylactic measures. If, after lowering the gilt to the horizontal, the vacuum was formed but no fluid was recovered, the gilt was returned to the laparoscopic position, the tubing was checked to make sure it was still in position, then an addi- tional 6 ml. of saline were injected and collection attempted while observing through the laparoscope. Three attempts were made to attach a cannula through the skin into the uterus. It was hoped that by inserting several centimeters of tubing into the uterine horn it would remain in place without being attached to the uterus. In the first attempt the cannula was attached to the skin just inside the left leg of the animal with a purse-string suture. The gilt was then housed by herself in a recovery pen. She worked the tubing out within three hours of re- covery from anesthesia. In a second attempt tag cement was used to attach the catheter to the outer body wall. Xylocaine (2%) was 27 injected subcutaneously to prevent irritation and rubbing. Once again the gilt was housed by herself. The tubing remained in place overnight, but within 24 hours, the tubing had worked out. In the third attempt, tag cement was again used, a piece of plastic and a gauze patch were placed over the cement. It was felt that the previous gilt had worked out the tubing by rubbing the tag cement, which was sticking to objects allowing her to pull out the tubing. Once again, within 24 hours the tubing had worked out. For a successful chronic cannulation the gilt should be restrained in some method to prevent her rubbing the cannula. In order to keep the cannula in place a plastic implant attached to the cannula and sutured under the skin may prevent the gilt from pulling it out. RESULTS Weight Gain The average and the range for the breeding weight, gain from days 1-30, 30-70, 70-110, total gain during gesta- tion and litter size are shown in Table I. The litter size and breeding weight for all the sows, divided into parity, is given in TableIEL The weight of the dam at breeding had no effect on litter size when breed and parity differences were removed. This was true for all parities. Parity 3 had the largest number of sows and the breeding weight ranged from 278 1b to 395 lb. The largest litter size from parity 3 sows was 14 and the smallest litter size was 5. The body weight gains of the dam from days 30 through 70 and 70 through 110 of gestation were not significantly associated with changes in litter size. However, the weight gain from days 1 to 30 showed a slight trend (p‘10.l7) toward increased litter size as the weight gained increased. Figure 2 shows the graph of the weight gain of all sows through 110 days of gestation. Not all sows were weighed on the same day of gestation so weights were interpolated for uniformity. Weight gains were constant throughout gestation when weights were taken at approximately 2 week 28 29 .mocsom CH cm>flm mum munmflm3 dads os.sm Hq.maa and o.mm chow sauce mm.~a ms.m~ o.mm o.H- oaauos mama came NG.HH mv.mm o.mo 0.0H onuom mama came mm.ma ma.mm o.so o.m omna mama came m.m mm.m o.mH 0.4 muwm umuuflq om.mv sa.mmm ass mum .unmfimz HmflufiaH soHp0fl>mQ cam: msam> msHm> wuwccmum ESEHXME ESEHCHZ H usmfiflummxm so mzom mv How munmflmz mmmnm>4 was EdEHGHZ .EDEHNMS .H magma 30 Table II. Litter Size, Breeding Weight, Weight Gain During the First 30 Days of Gestation, and Total Weight Gain During Gestation, Listed According to Litter Size and Parity Litter Breeding Gain During Total Size Weight Days 1-30 Gain Parity 2 4 289 29 131 4 388 30 120 5 293 41 147 8 316 28 138 9 305 27 115 10 293 45 144 10 298 56 162 10 319 26 149 10 349 29 95 11 294 55 138 11 314 64 116 11 314 52 128 13 286 44 154 Parity 3 5 278 45 119 6 362 36 146 6 367 34 98 8 349 21 109 9 344 34 134 9 354 51 134 10 314 42 131 10 330 41 150 10 374 46 126 11 318 48 145 12 296 52 179 12 395 43 117 13 306 56 129 13 329 43 125 14 335 45 111 Parity 4 8 371 29 39 10 421 30 56 12 398 17 84 14 407 3 41 15 363 41 143 continued 31 Table II--continued Litter Breeding Gain During Total Size Weight Days 1-30 Gain Parityg5 and Greater 7 405 28 101 8 355 39 126 9 348 27 120 9 352 22 91 9 372 4 43 9 414 24 82 9 426 21 94 9 447 23 73 10 368 47 148 14 448 25 50 15 435 14 57 15 444 41 122 32 $4 5 .mossom a.“ sofiumummm mcausv m30m mo saw-m ”:3me mmmum>¢ .m musmflm 203.3.me mo >40 0: oo. om 00 Q. om on o¢ Om ON 0. can Ohm 0mm o; 09. 00¢ 05¢ omv 1H913M 33 intervals. The sows were graphed according to breed and parity, but the slope of the curves were similar, so only the graph for all sows is included. Table III shows the differences in litter size and weight gains during gestation between the breeds and pari- ties used in the experiment. The weight gain of the sows in parity 2 or 3 was significantly more than the gain of sows in parity 4 (p‘<0.001). The differences between litter sizes from different parity sows was not significant. (Experiment 2) Figures 3 and 4 indicate the relationship between body weight changes during the first 24 days of gestation and plasma progesterone levels. In order to standardize the different initial weights of the sows, the graph was made according to percentage of initial body weight at day 3 of gestation. The weights recorded and their percentages are listed in Tables IV, V and VI. Due to the limited number of sows and the variability of individual animals statistical significance could not be attached to the weight changes. However, it is interesting to note that the weight gains do not rise consistently during this period but rather peak at day 15 then decrease. This seems to follow the progesterone pattern during this early part of gestation. Progesterone peaks at day 12 then decreases (Figure 3). Increasing the progesterone levels on days 15-20 causes a modest upswing of 34 mm.oa m.HH hm.m mm.m mNHm Hmuuflq mmmum>¢ om.~m ov.mn hm.ama No.mma soaumummw mcwuso CHMO Hmuoe mmouu sm.m ms.mma G H 0H m muflnmmsmm Imnflnmxnow mm.aa mm.om v H N H muflnmmsmm mn.m mv.nm m m m w muflnmxnow muwm kuuflq sflmo who: no mmmuw>¢ mmmnm>< m huflumm v mpfiumm m wuflumm N muflumm mzom mo Hmufidz auflnmm was cmmnm ou mcflpuooofi cmumfiq mNflm kupwq was mzom mo :Hmw usmfimz mmmnm>¢ .HHH OHQMB 35 mo mm >80 :0 msflccm can ma mmo mcflscwmwm mmmo OH How maouummuwcoumumwmoum saws pmuomncH msom mo unmwmz m man no mommusmoumm paw munmfimz >Uom vo.a mm.o Hm.o Ho.H hm.o no.0 uonum pumvswum mm.m mo.m mm.m ov.m Hm.o mv.H cofiuMA>mQ pumocmum v.~oa m.moa n.moa mv.m0H m.Hoa m.ooa cam: o.moa m.moH N.moa m.moa m.HOH v.mm ooa .uz m man m0 w mmm @mm vmm mmm 55m mom mum .u3 Hmsuom mlhaa m.~oa @.~oa m.NOH «.moa o.HoH m.moa ooa .uz m wmo m0 m vmm mom vmm 0mm 5mm mmm mmm .uz Hmsuom Hauvom o.voa o.moa m.moa o.voa o.HOH m.aoa ooa .uz m hmo m0 w mmm mum mmm mmm whm mum mom .uz Hmsuoa mlvoa m.voa m.voa m.moa m.ooa o.Hoa m.ooa 00H .uz m man m0 m «av vav mav mmv How oov 5mm .uz Hmsuod HTNH o.nm m.mm n.mm m.mm @.Noa m.mm ooa .uz m mmo m0 w omv oov How Neg mhv mow new .uz Hmsuom mlbm h.HOH m.Hoa o.moa m.moa «.moH h.aoa ooa .uz m Mme m0 w oav mav haw mmv mmv oav mow .uz Hmsuom mlhm mm ma ha ma HA 5 m Hmnadz meo Hmaflsd cofiumumow .>H OHQMB 36 oa.H No.0 Hm.o No.0 mo.H mo.H no.0 Houum pumpsmum >.N mm.m vm.m ao.m mm.m mm.m mm.H QOHHMH>mo oumcsmum m.aoa H.moa h.moa o.m0H Hm.m0H m.moa m.aoa cmwz m.mm m.mm m.mm m.ooa N.ooa v.aoa ooa ooa .pz m man m0 w Hmv mmv Hmv «mv ,mmv mmv «mg «mg .uz Hmsuod Hannmm m.ooa H.Hoa S.Hoa >.Hoa m.Hoa m.wm m.ooa ooa .uz m use m0 w com 50m mom mom onm mmv vmm mom .uz Hmsuod mind h.¢oa v.voa H.voa h.voa v.moa m.moH m.NoH ooa .uz m mmo m0 w mow vow mov mow mov Hov mom 5mm .uz Hmsuom vtvom v.voa m.voa m.moa m.moa m.moa ¢.voa v.voa ooa .uz m >80 mo w wow wow wow oov wow Nov Nov mom .uz Hmsuod mnhoa p.50 m.ooa m.ooa m.aoa o.aoa o.aoa m.aoa ooa .uz m man no a vmm vmm omm mam mom 5mm mmm mmm .u3 Hmsuom mum m.Hoa m.m0H m.v0H o.voa o.ooa o.ooa h.moa ooa .93 m use m0 w mHv mav adv mav mmv omv mag Nov .ux Hmsuom mlmoa mm Hm ma ha ma HA 5 m Hmnfisz HmEHad beam mBOm Houusoo mo usmflmz m use mo mmmucmoumm was munmflmz moon .> magma mo.H mm.o om.o mh.o mh.o Hm.o HOHHM Unmvcmum 37 v.m mm.H mH.~ wh.H hm.H MH.H coHuMH>mo oumpcmum S.HOH m.HOH m.OOH H.HOH m.HOH m.HOH com: nmunm usosuH3« mn.H Hm.H mm.H mn.o v.H mo.o Houum oumocmum mm.v mm.m o.m m.H vv.m n.H coHumH>mo unaccmum m.00H H.00H o.ooH n.00H m.00H mm.HOH cmmz o.mm o.mm m.vm m.mm H.5m m.mm 00H .uz m hmo mo w mum mum omm mow mom vow HHv .uz Hmsuod mnnm« m.v0H m.m0H m.m0H m.HOH m.HOH m.HOH ooH .uz m Smo mo w mmm mmm mmm omm mmm 0mm va .uz Hmsuod HHImOH m.vOH m.moH n.NOH m.HOH m.00H m.HOH 00H .u: m amo m0 w omm 0mm vmm Hmm hum mum mum .uz Hmsuog ounmm n.mm OOH n.mm n.mm m.OOH m.00H OOH .uz m >80 no a 00m Hmm omm omm vmm vmm Hmm .uz Hmsuom Nunm m.NOH m.m0H m.m0H m.m0H m.moH m.mOH 00H .uz m moo m0 w omm omm mmm mmm mmm mmm mmm .uz Hmsuod mIHHH m.mm m.mm mm m.mm m.mm o.HOH 00H .uz m mmo m0 a mmm mmm omm nmm mom wow wow .u: Hmsuog mummm m.00H o.NOH v.mm n.HOH m.~0H m.m0H 00H .uz m use mo w omm vmm mvm mmm mmm 0mm nvm .uz Hmsuod OHTv vm Hm mH mH NH m w m Honssz mama HMEHcm UmHm uoz mHmEHcm Houucoo wo uanmS m mmo mo mmmucmoumm was mugmHmB Hmsuog .H> mHnma 38 NG/ML PROGESTERONE 0— ON on 3 mm zv .UmHo m30m Houusou zo_h<._.wu¢ no >40 ON . n. O. .m musmHm ..w>u._ wzommhmmoomm T .. n... 1. Flo—w; 3.1111 .0. NO. no. «.0.— no. mo. 1H913M '°/o 39 Am zv .msouumw\mcoumummmoum zuHB UmuomflsH msom .v musmHm 29. keg. Wm o no >40 mN ON 0. O. n .m musmHm OO. _O_ NO. MO— VO— no. mo. 95 lHSEN“ 41 the weight curve (Figure 4). The body weight gain during the first 25 days of gestation of the six control sows not bled is shown in Figure 5. A seventh control sow was weighed but she consistently lost weight during this 25 day period. This is different than any other sow weighed in the experiment. The possibility that this sow may have been sick cannot be dismissed. Therefore, the sow's weight is included in Table VI but not on the graph. Table VI gives the % weight for the days with and without this sow. The weights in the control sows peaked earlier than the sows which were bled. Uterine Fluid The recovery of uterine fluid by laparoscopy has been attempted forty-six times and uterine fluid has been re- covered thirty-six times. The total recovery of fluid flushed into the uterus was 32% including those attempts which were unsuccessful. Unsuccessful attempts were due to a lack of pooling of the fluid in the uterus. As a result the saline tended to flow away from the inserted tubing. In the attempts which were successful, the recovery rate was 40%. These results are shown on the following page. 42 Number of Saline Amount Fluid Percentage Injections Injected Recovered Recovery Total attempts 46 272.8 87.65 32% Successful 36 221.8 87.65 40% The insertion of the polyethylene tubing down the 15 ga. needle proved to be the most successful method tried. Initially attempts were made without blocking the end of the tube, but no fluid was recovered. The saline was injected too far away from the end of the tubing making pooling around the opening unlikely. At times the tubing slipped out in the process of lower— ing the gilt. This was detected by the absence of a vacuum while attempting to aspirate fluid from the uterus. In these cases, the gilt was returned to the 30° angle, and the tubing was reinserted usually in the same uterine horn. If, after lowering the gilt to the horizontal, the vacuum was formed but no fluid was recovered, the gilt was returned to the laparoscopic position, the tubing was checked to make sure it was still in position, then an addi- tional 6 ml. of saline were injected and collection attempted while observing through the laparoscope. DISCUSSION The weight at breeding of the sow had no effect on litter size when breed and parity differences were removed. In Table II the breeding weight and the litter size are listed according to parity. From this data there appears to be no relationship between litter size and parity. This is in contrast to Heap gt_gt. (1967) who increased litter size by 4.2 offspring with an increase in breeding weight to 100 1b. However, Heap was slaughtering the sows at day 28 of gestation and it is possible that if the sows had completed gestation the increase may not have been as great. The weight gains from days 30 to 70 and 70 to 100 proved not to be significantly correlated with litter size, but the weight gain from days 1 to 30 showed a slight trend (p‘<0.l7) toward increased litter size as gain increased. These values are also given in Table II. The data was analyzed by least squares regression to remove the effects of breed and parity. This trend toward larger litters is in agreement with McKenzie (1928) who showed increased litter size with sows showing increased gain during the first four weeks of gestation. During the first 30 days of gestation, the fetus weighs very little, reaching a weight 43 44 of 1.5 gm. at day 30. The majority of the fetal weight gain occurs towards the end of gestation (Ullrey gt gt., 1965). Fetal weight gain alone cannot account for the weight causing the trend in increased litter size. Some other factor is increasing the weight and embryonic survival. The ability of steroids and related compounds to affect weight gain is well documented with implants of diethyl- stilbestrol (estrogen activity) causing increased feed efficiency in feedlot cattle. In the pig, Salmon-Legagneur and Rerat (1962) reported the ability of the pregnant sow to gain on what would be a maintenance ration for a non- pregnant animal. The possibility that progesterone combined with small amounts of estrone may be causing these weight changes was studied in six pigs which were weighed and bled during early gestation. As can be seen from Figure 3, there appears to be a correlation between the changes in body weight and progesterone levels with the weight peaking at day 15 and progesterone at day 12. Reddy gt gt. (1958) us— ing a 2000:l progesterone-estrone ratio had a significant increase in the weight of the uterus after the embryo had been removed at day 56 of pregnancy. The uterus of the gilts treated for a 10 day period with 25 mg. progesterone and 12.5 micrograms estrone starting at day 13 of gestation, weighed 15.74 lb. Assuming 1 ml. of fetal fluids weighs 0.0022 lb. (weight of 1 ml. of water), the weight of the 45 uterus was 7.12 lbs., less the fetal fluids. The control group uteri weighed 10.57 and 4.15 lbs. respectively. Progesterone-estrone is shown here to affect the weight of the uterus at day 56 and it is plausible that it is affect- ing uterine weight during early gestation while it is being administered. Six sows were given progesterone-estrone injections which served to increase plasma progesterone levels. These elevated levels did not prevent the drop in weight after day 15, but the drop was not as great as that observed in the bled controls (Figure 4). Daily fluctuations in weight are usually associated with changes in fluid volume of the body. Progesterone is known to affect uterine secretions and for a maximum secretion small amounts of estrogen must be used in conjunction with the progesterone (Knight gt gt., 1974). Thus, part of this weight fluctuation may be accounted for by changes in the fluid volume of the uterus. The control sows which were not bled (Figure 5) showed a drop in weight but this occurred earlier than in the two bled groups. The total gain during gestation did not affect litter size when breed, parity and boar differences were removed. This disagrees with Buitrago gt gt. (1974) who found a sig- nificant and positive correlation between gestation gain and litter size. This may indicate the difference between 46 gilts and sows, as Buitrago used gilts. An overall gain in weight reflects a better utilization of nutrients for a gilt which is still growing, compared to a sow which has already matured or is growing at a slower rate. Thus, large weight gain in the gilt may indicate good condition while the same amount of gain in the sow would indicate overfeed- ing and poor condition. Young gt gt. (1974) showed a posi- tive and significant correlation between average daily gain and ovulation rate with gilts. Thus, if faster growing gilts have more ovulations, this may be reflected in larger litters. However, second and third parity sows are still growing as indicated by the additional amount of weight gained during gestation when compared to fourth parity sows. These sows showed no indication of gain during gestation being related to litter size (Table II). As was just noted, sows in parities 2 and 3 gained significantly more than sows in parity 4 or greater. However, the higher parities averaged larger litters. This indicates that even though they are no longer gilts, sows at parity 2 or 3 are still growing, and thus utilizing more feed for growth purposes. The larger litter size in the older sows could be a reflection of less competition between the sow and her litter for nutrients. Increasing litter size with increased parity has been reported before by Olbrycht (1943) and Korkman (1947). The trend is for litter 47 size to increase to a peak after the first few litters then slowly decrease. This trend was found with our sows as litter size increased through parity 4 then decreased slightly (Table II). Even though statistical significance was not found in litter sizes between the different parities the difference could be real. Reddy gt gt. (1958), showed a significant increase in litter size, measured at day 55, when injections of 25 mg. progesterone and 12.5 micrograms estrone were administered for a 10 day period starting on day 13 of gestation. These results have been reproduced recently by Wildt (1975) who narrowed the time of injection to days 16 and 17 of gesta- tion and still produced a significant increase in litter size. Thus, it is obvious that the hormone injections are modifying the uterine environment in some way to cause a decrease in embryonic mortality. Furthermore, it takes a synergistic action between progesterone and estrone to cause the effect. The protein content of the uterine fluid changes depending upon the stage of the estrus cycle. It increases rapidly at day 12, peaks at day 15, and decreases very rapidly to a low level by day 17 (Murray gt gt., 1972). The decrease in protein coincides with a decrease in pro- gesterone as shown by Guthrie gt gt. (1972). Knight gt gt. 48 (1973) showed that total protein content was maximum when a progesterone-estrogen combination was used on ovarectomized gilts. The five protein fractions described by Murray _t~gt. (1972) were present in the progesterone-estrogen group, but in the group just given estrogen, only fractions I, II, and III were present. From the above work, it can be seen that uterine pro- teins are influenced by estrogen and progesterone, and it is possible that the increased embryo survival found by the administration of these two hormones during the period of implantation may be mediated by these proteins. The usefulness of a procedure for the collection of uterine fluid by laparoscopy is readily apparent for it allows the collection of protein without the trauma of laparotomy. This makes possible the study of protein changes within the uterus of individual animals during the estrous cycle and early pregnancy. SUMMARY AND CONCLUS ION Forty-six sows were weighed regularly throughout gestation and analyzed using a least-squares regression for weight gains correlation with litter siZe. Twelve addi- tional sows were bled and weighed through the first twenty- five days of gestation. Six of these sows were injected with progesterone:estrone to elevate the progesterone levels. Weight gains had little correlation with litter size except during the first thirty days when a trend toward increased litter size was evident with increased gains. Closer inspection of this initial period showed weights, rather than increasing gradually, peak approximately day 15 then decrease. This rise then fall is also evident in peripheral plasma progesterone levels, which peak at day 12 then decrease. Increasing the plasma progesterone levels with injec- tions of progesterone:estrone from days 13-22 of gestation did not serve to stop this loss in weight but the drop was less than the control-bled sows. A method was also introduced for routine collection of uterine flushings utilizing laparoscopy. Seventy-nine 49 50 percent of our attempts were successful and forty percent of the fluid injected was recovered in the successful attempts. LITERATURE CITED LITERATURE CITED Adams, C. R., D. 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R. Dukelow. Submitted to J. Reprod. & Fert., November 1975. Abstracts Presented at Midwestern Section of the American Society of Animal Science. 1 COLLECTION OF OVA and UTERINE SECRETIONS BY LAPAROSCOPY C. B. Morcum, David E. Wildt and W. R. Dukelow Michigan State University, East Lansing The use of the laparoscope for in yizg_observation of reproductive phenomena in swine has been previously reported by our laboratory. This paper deals with two techniques for the laparoscopic collection of ova and uterine secretions. Ova were collected with a 22 gauge 4" needle inserted via a cannula through the abdominal wall. The ovarian ligament is grasped with laparoscopic forceps and the needle is then directed into follicle and the contents aspirated by vacuum from a syringe. The follicular contents are then examined 60 61 under a dissecting microscope for ova. To collect urterine secretions a 16 gauge 1%" needle is inserted through the abdominal wall. The uterine horn is grasped with laparoscopic forceps and brought up to the ventral abdominal wall. The needle is directed into the uterine lumen. PE90 tubing (with the end sealed and perfor- ations along the distal axis) is then inserted through the needle such that all the openings are intrauterine. The uterine horn is allowed to return to its normal position. Ten milliliters of 0.15M NaCl are injected into the lumen and then aspirated into a syringe. These two procedures allow the collection of samples without the major stress of surgery and permits the monitor- ing of a single animal over an extended period of time. VITA Name: Clifford B. Morcom III Born: August 5, 1950 Place of Birth: New York, New York Formal Education: Monmouth Regional High School New Shressbury, New Jersey Pennsylvania State University State College, Pennsylvania Degrees Received: Bachelor of Science, Pennsylvania State University 1972 Member of: Phi Sigma Gamma Sigma Delta 62 MICHIGAN STATE UNIVERSITY LIBRARIES I III ll'llllll II Illll I . O1 3 1293 3 42 8836