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The Technique of Electroejaculation and Its Use in Dairy Bulls1
T H E T E C H N I Q U E OF E L E C T R O E J A C U L A T I O N ITS USE IN DAIRY BULLS 1
AND
P. J. DZIUK, E. F. GRAHAM, AN]) W. E. PETERSEN
Departv~ent of Dairy Husband~'y University of Minnesota, St. 2aul The widespread use of artificial insemination has increased the value of each ejaculate of semen f r o m valuable sires. Since some males are either unwilling or unable to serve an artificial Vagina, their usefulness is limited. Electrical stimulation is one of several methods that have been used in an a t t e m p t to collect semen f r o m such animals. One of the first to record the obtaining of semen b y the use of electrical stimulus was Batelli (1), who collected f r o m guinea pigs. Moore and Gallagher (15) modified the technique and used it in an evaluation of the functioning of the accessory glands of the guinea pig. G u n n (5) was the first to make extensive use of electrical stimulus as a means of collecting semen f r o m rams. F u r t h e r work with this technique was done b y Bonadonna (2), L a m b e r t and 1VicKenzie (6), Terrill (17), and others. The stimulation was 60 cycle a l t e r n a t i n g current of about 30 volts applied b y means of a rectal probe and a l u m b a r electrode. L a p l a u d and Cassou (7) constructed a bipolar rectal probe which p e r m i t t e d stimulation without an external l u m b a r electrode. L a u r a n s and Clement (10), Likar and K a m h i (11), Mata and Cano (14), and O r t a v a n t et al. (16) made f u r t h e r use of the bipolar electrode in collecting semen f r o m rams. The bipolar probe was a d a p t e d b y Thibault et al. (18) so t h a t it could be used in the stimulation of bulls. His equipment consisted of a probe with 30 brass rings insulated f r o m each other by ebony. H e used 60 cycle a l t e r n a t i n g c u r r e n t which was varied f r o m zero to t h i r t y volts and back to zero every 3 to 5 seconds. Evidence is given on p i t , volume, density, and motility of the semen to indicate t h a t the semen samples collected were normal. Thibault (19) described i~l detail the electrical circuits, construction of the probe, and the technique used in r e s t r a i n i n g the bull and collecting the semen. L a p l a u d e t al. (9) f u r t h e r elaborated on the application of electrical stimulus as a method of obtaining semen f r o m bulls. L a p l a n d and Cassou (8) cite an example of a young bull that had yielded 161 ejaculates in an artificial insemination center before an accident which affected his h i n d q u a r t e r s p r e v e n t e d a n y a t t e m p t at service. A 6-ml. sample of semen was obtained b y bipolar electrical stimulation and used to inseminate two cows. A pair of f r a t e r n a l twins and a single calf were b o r n f r o m these matings. L u t w a k - M a n n and Rowson (12) made several chemical determinations of various fractions of bull semen collected b y electrical stimulus. They found t h a t the p r e - s p e r m fraction was not f r o m Received for publication February 1, 1954. 1University of Minnesota Scientific Journal Series Paper No. 3108, i~innesota Agricultural :Experiment Station. 1035
1036
e - J. D Z I U K E T AL
the accessory glands but p r o b a b l y f r o m the urethral glands. Excesses of this fluid did not affect motility or fructolysis. E g r o n (4) f o u n d the semen collected by means of artificial vagina did not differ greatly f r o m t h a t obtained b y electrical stimulus. The p t I was slightly higher and the density of s p e r m was lower in semen resulting f r o m electrical stimulation. Teasing the bull before collection increased the density. Briere (3) mentioned the applications of eleetroejaculation in his thesis. Mascarenhas and Gomes (13) were the first to report large numbers of collections by electrical stimulation in the bull. They used the multi-ringed bipolar probe to a p p l y both constant and impulse type of current. The impulse type of c u r r e n t is described in which the stimulation is passed f r o m one group of rings to another in r a p i d succession. This type of stimulation gave less fluid and the side reactions were more marked than with the constant type. Initial experiments were p e r f o r m e d by using a 15-v. current, b u t later work was done with a current of 5-10 v., which also proved satisfactory. A total of 48 separate collections were made f r o m seven bulls. Generally, the semen was less dense but of higher volume so that the total numbers of sperm were essentially the same as in semen collections made with an artificial vagina. E X P E R I M E N T A L PROCEDURE
Eighteen bulls were used as experimental animals in these trials. Holstein, Jersey, Guernsey, Brown Swiss, and Shorthorn breeds were represented and, except for a set of f r a t e r n a l Holstein twins, all were p u r e b r e d animals. Several different types of electrical stimuli were tried with v a r y i n g degrees of success. The o r d i n a r y inductorium stimulus of high voltage peaks and low a m p e r a g e was found to be highly stimulating to motor nerves, especially the sciatic, but was ineffective in causing either erection or ejaculation. The frequency of the stimulating current was varied f r o m 15 to 900 cycles per second b y means of a variable frequency oscillator. No advantage was found for either the higher or lower frequencies. Because of the expense and inconvenience of the equipment, s t a n d a r d 60-cycle current was used subsequently. A 30-v., 60-cycle alternating current which could be varied f r o m zero to 30 v. by means of a rheostat also was tried. Because of undesirable fluctuations in the voltage applied to the bull as the c u r r e n t load increased, the rheostat was discarded in favor of a variable t r a n s f o r m e r as a means of v a r y i n g the voltage level. An isolation step-down t r a n s f o r m e r was found to be necessary in isolating the circuit from the power source and to allow more gradual changes in voltage. A t t e m p t s were made to include resistors v a r y i n g from 25 to 200 ohms in series with the bull in the final circuit. This was done to reduce marked changes in the current flow through the bull as the resistance changed because of feces accumulation and poor contact. Some mechanism, p e r h a p s contraction of the rectum upon stimulation, causes the effective resistance of the bull to decrease as the a m p e r a g e flow increases. Therefore, the added series resistor prevented a sufficiently high amperage level to achieve either an erection or an ejaculation.
ELECTROEJACULATION IN DAIRY BULLS
1037
Several rectal probes of various dimensious and specifications were used in these trials. The early a t t e m p t s were made with only two electrodes. These were moved and placed b y insertion of the gloved a r m into the rectum. B y localizing the stimulation over the glands and o v e r the bulbo-cavernosus muscle, pronounced erection and a v e r y little fluid could be obtained; however, in no case was an ejaculatiou obtained. One probe 1 in. in diameter with seven rings 13A in. a p a r t was used with some success but did not give consistent results. A second probe, 1 in. in diameter with nine rings 11A in. apart, gave much more fluid, but ejaculation did not follow in all cases. A smaller probe, 3/~ in. in diameter with six rings 3A in. apart, gave v e r y little stimulation even with the resulting high amperage. A probe 2 in. in diameter with three longitudinal electrodes was used but was found to be less desirable t h a n the ringed probe of similar dimensions, because of more side reactions and more fluid secretion. The probe which seemed most satisfactory was 11/2 in. in diameter with six rings 13~ in. apart. This probe was made f r o m a section of 11/2 in. h e a v y steam hose 2 feet long. E a c h ring electrode was p a r t i a l l y inlaid into the rubber. The electrodes were made f r o m 2 turns of h e a v y solid core solder. A separate lead r a n inside the probe and was soldered to each electrode. Most of the results reported were obtained with this t y p e of probe and the electrical circuit shown in F i g u r e 1. II0/30-Volt transformer
~,,2-Amp fuse ~
\ \ \ \ " \ "--Ammeter ' ~_ ~
22"
>1
,oT¢ l
~Voltmeter Rectal probe ~-Varioble transformer
FIG. 1. Schematic diagram of stimulator. Collection of the semen was made b y means of a r u b b e r cone which was held open by a section of radiator hose 11/~ in. long, similar to t h a t used for the artificial vagina. The opening was protected fronl dirt b y a stiff r u b b e r sheet with several intersecting slits cut in it. The semen was collected in a test tube which was s u r r o u n d e d b y a larger test tube to serve as a w a r m w a t e r jacket to minimize cold shock. I t is necessary to be able to see the semen at collection to aid in deciding on the n u m b e r and intensity of subsequent stimulations. The technique used became f a i r l y well established as the trials progressed. The first step is administration of an enema of about two quarts of a w a r m 5% solution of sodium chloride. I t is i m p o r t a n t to give an adequate enema to remove as much of the fecal material as possible in order to minimize its insulation of the electrodes. The p r e p u t i a l area should be clipped and brushed to reduce the amount of d i r t and foreign material which otherwise m a y get in the semen. The probe is lightly lubricated with some noninsulating material a n d introduced into the rectum. The probe is inserted f o r v a r y i n g distances,
1038
P. $. D Z I U K ET AL
d e p e n d i n g on the size of the bull, b u t g e n e r a l l y no more t h a n 12 in. The best results, especially as f a r as e r e c t i o n is concerned, were g e n e r a l l y o b t a i n e d w h e n the p r o b e was m a i n t a i n e d on the v e n t r a l side of the r e c t u m . The voltage was g r a d u a l l y i n c r e a s e d to a b o u t 3-5 v. a n d t h e n back to zero e v e r y 5-10 seconds. The s u b s e q u e n t s t i m u l a t i o n s go p r o g r e s s i v e l y h i g h e r so t h a t at a b o u t the fifth s t i m u l a t i o n the m a x i m u m of 10-15 volts is reached. There is m u c h v a r i a t i o n b e t w e e n bulls, a n d f r o m one time to a n o t h e r w i t h the same bull, so t h a t e x p e r i e n c e best g o v e r n s the r a p i d i t y of increase. E r e c t i o n a n d fluid secretion u s u a l l y s t a r t at a b o u t the 5:v. level, b u t e j a c u l a t i o n u s u a l l y occurs at the 10-15 v. m a x i m u m , a t which time a b o u t 0.5 to 1.0 a m p e r e of c u r r e n t is flowing. Too m a n y s t i m u l a t i o n s a t a s u b - e j a c u l a t o r y level y i e l d m u c h fluid a n d o f t e n make i t difficult to o b t a i n a n e j a c u l a t i o n at a n y level. Too r a p i d a n increase m a y lead to a p r e - e r e c t i o n e j a c u l a t i o n a n d c o n t a m i n a t i o n of the semen w i t h dirt, since the p e n i s is n o t free of the sheath. A t the time of s t i m u l a t i o n m a n y m o t o r n e r v e s are also s t i m u l a t e d , a n d g e n e r a l t e t a n y of the h i n d legs results. T h e r e is a g e n e r a l e x t e n s i o n of the h i n d legs a n d a t e n d e n c y to p u s h f o r w a r d . However, i n n o ease has a b u l l been observed to establish u n f a v o r a b l e associations w i t h e i t h e r the collection stall or the m e t h o d of collection. I t was f o u n d t h a t a m i n i m u m of r e s t r a i n t was n e c e s s a r y f o r best collection, the o n l y r e q u i r e m e n t b e i n g a s t u r d y s t a n c h i o n or stock to r e s t r a i n the neck a n d to allow the b u l l to p u s h f o r w a r d , Good footing, such as e a r t h or gravel, was needed, since the b u l l s s l i p p e d on concrete or wooden floors. TABLE 1 Collection data
Bull No.
No. of successful electroejaculations
:No. of attempts
Total volume of fractions containing sperm
Average volume
1 2 3 4 5
29 2 3 0 1
30 3 4 3 1
(ml.) 327 33 30 0 10
(ml.) 11 i6 10 0 10
6
6
6
74
12
7 8 9 10 11 12 13 14 15 16 17 18 19
9 13 9 12 12 2 3 1 2 2 3 1 1
10 13 10 12 12 2 3 1 2 2 3 1 1
93 189 216 95 77 32 36 9 33 39 63 8 11
10 14 24 8 6 16 12 9 16 19 21 8 11
Total
111
119
1,375
ELECTROEJACULATION IN DAIRY BULLS
1039
RESULTS AND DISCUSSION
One h u n d r e d eleven s e p a r a t e collections were m a d e f r o m 18 d i f f e r e n t bulls. S e m e n was n o t o b t a i n e d in e i g h t a t t e m p t s . Of t h e 19 b u l l s used, o n l y one f a i l e d to y i e l d a n y semen. T h r e e s e p a r a t e a t t e m p t s w e r e m a d e w i t h t h i s bull, b u t d e a t h d u e to p n e u m o n i a p r e v e n t e d f u r t h e r a t t e m p t s . T h e v o l u m e of s e m e n o f t h e 111 s u c c e s s f u l collections a v e r a g e d 12.3 ml. w i t h a r a n g e of f r o m 1 to 35 ml. ( T a b l e 1). The d e n s i t y r a n g e d f r o m v e r y low to 1.27 b i l l i o n p e r m i l l i l i t e r i n a l m o s t a n i n v e r s e r e l a t i o n s h i p to volume. T h e e x t e n t of s t i m u l a t i o n a n d d e g r e e of s e p a r a t i o n a r e t h e g o v e r n i n g f a c t o r s of t h e d e n s i t y - v o l u m e r e l a t i o n s h i p . U s u a l l y , no a t t e m p t was m a d e to f r a c t i o n a t e t h e fluid p o r t i o n s f r o m t h e m o r e dense semen, since it was b e l i e v e d t h a t the excess fluid was n o t so d e t r i m e n t a l to t h e s e m e n as was t h e n e c e s s a r y i n t e r r u p t i o n i n t h e sequence of collection w h e n t h e f r a c t i o n s were s e p a r a t e d . O f t h e u n s e l e c t e d s a m p l e s in w h i c h t h e t o t a l n u m b e r of s p e r m was d e t e r m i n e d , t h e r e was no a p p r e c i a b l e difference i n t o t a l n u m b e r s of s p e r m b e t w e e n s a m p l e s collected b y m e a n s of a r t i f i c i a l v a g i n a a n d those collected b y e l e c t r i c a l s t i m u l a t i o n . A l t h o u g h t h e r e w e r e i n d i v i d u a l b u l l differences, t h e first few s t i n m l a t i o n s u s u a l l y y i e l d e d some fluid, f o l l o w e d b y t h e m o r e c o n c e n t r a t e d semen. F u r t h e r s t i m u l a t i o n y i e l d e d m o r e d i l u t e s e m e n a n d f i n a l l y c l e a r fluid. One n o r m a l b u l l was collected f r o m b y e l e c t r i c a l s t i m u l u s a b o u t once w e e k l y f o r a t o t a l of 30 c o l l e c t i o n s ( T a b l e 2). H e f o r m e d no u n TABLE 2 Evaluation of unselected semen samples collected by electroejaculation from buU No. 2
Collection :No.
Total volume
Total sperm :No. in billions
(ml.) 1 2 3 4 5 6 7 8 9 10 a 11 12 13 14
5 9 18 31 13 5 4 15 12 10 7 20 5 11
Motility rate (%)
5.0 5.7 7.0 15.5 16.4 7.6 1.2 10.3 9.6 10.5 6.4 21.0 4.0 13.4
85 85 95 85 85 85 80 85 75 95 80 85 50 80
a Collected by artificial vagina 5 minutes after collection :No. 9. f a v o r a b l e a s s o c i a t i o n w i t h the t r e a t m e n t a n d was n o t affected in a n y o t h e r w a y . H e did, on s e v e r a l occasions, serve a n a r t i f i c i a l v a g i n a w i t h i n a f e w m i n u t e s a f t e r collection b y e l e c t r o e j a c u l a t i o n a n d p r o d u c e d a n o r m a l e j a c u l a t e . N i n e of t h e b u l l s i n a r t i f i c i a l i n s e m i n a t i o n s t u d s were i m p o s s i b l e to collect f r o m w i t h t h e a r t i f i c i a l v a g i n a . F o r t y - s e v e n collections w e r e m a d e f r o m these bulls. T w e n t y - s e v e n of these collections were d i l u t e d a t u s u a l r a t e s a n d u s e d in t h e field d u r i n g t h e m o n t h s of M a r c h , A p r i l , M a y , a n d N o v e m b e r , 1953. R e s u l t s
P . J. D Z I U K
1040
ET AL
TABLE 3 Conception data on bulls which were electroejaculated
Bull
A
B
C
D
Number of first services 60-90-day nonreturns ( % ) Previous year over-all ( % ) 60-90 day nonreturns
28 68.0 67.0
2,012 60.4 61.9
138 44.9 55.0
89 57.3 61.5
o f t h i s t r i a l a r e s h o w n i n T a b l e 3. T h e c o n c e p t i o n d a t a f o r t h e r e m a i n i n g c o l l e c t i o n s is n o t a v a i l a b l e at t h e t i m e of w r i t i n g .
10
SUMMARY AND CONCLUSIONS E v i d e n c e h a s b e e n p r e s e n t e d w h i c h i n d i c a t e s t h a t i t is p o s s i b l e b y e l e c t r i c a l s t i n m l u s to c o l l e c t s e m e n f r o m s o m e b u l l s w h i c h a r e e i t h e r u n a b l e o r u n w i l l i n g to s e r v e a n a r t i f i c i a l v a g i n a . T h e r e h a v e b e e n n o a d v e r s e effects o r u n f a v o r a b l e a s s o c i a t i o n s i n b u l l s c o l l e c t e d f r o m b y e l e c t r o e j a c u l a t i o n . T h e s e m e n so o b t a i n e d w a s g e n e r a l l y of g r e a t e r v o l u m e b u t of l o w e r d e n s i t y . H o w e v e r , t h e t o t a l n u m b e r s of s p e r m a t o z o a w e r e c o m p a r a b l e to a n e j a c u l a t e o b t a i n e d b y a n a r t i f i c i a l vagina. E l e c t r o e j a c u l a t i o n m a y s e r v e as a v a l u a b l e a d j u n c t to t h e a r t i f i c i a l v a g i n a as a m e a n s of c o l l e c t i n g s e m e n i n a r t i f i c i a l i n s e m i n a t i o n c e n t e r s , e s p e c i a l l y f o r b u l l s w h i c h a r e d i f f i c u l t to c o l l e c t s e m e n f r o m b y m e a n s of t h e a r t i f i c i a l v a g i n a . ACKNOWLEDGMENT The authors wish to acknowledge the cooperation of John Dysart of Land O'Lakes Herd Improvement Division and Wallace Miller of Minnesota Valley Breeders' Association in making conception data available for this study. REFERENCES
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(6)
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F.
Une
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obtenir
l ~ Emission
Complete
du Liquide
des Vesicules
Seminales chez le Cobaye. Soc. de Physique et d' Hist. nat. de Gcneve, Co~np. Rend., 39: 73. 1922. BONADONNA,T. I1 Metodo dell' Elettroeiaculazione negli Ovini e nei Caprini. La Fccondazione Artificiale, 1: 54. 1938. (Supp. to Clin. Vet., 61. 1938.) BRIEI~E, J. Contribution a l'Etude des M@thodes de Collecte du Sperme chez le Taureau. Thesis 1-43 Alfort. 1950. (Abs.) Vet. Bull. 319'~'. 1952. EGR0~I, .... Sur quelques R~sultats Pratiques de 1' Electro-Ejaculation. Elevage ct Inse~n. No. 13: 19. 1952. Ani~na! Breed. Abstr., 21, No. 218. 1953. GVNN, ]~. M. C. Artificial Production of Seminal Ejaculation and the Characters of the Spermatozoa Contained Therein. Council for Scientific and Industrial Research, Bull. 94. 1936. LAMBEI~T~W. V., AND MCKENZIE, F. F. Artificial Insemination in Livestock Breeding. USDA Circular 567. 1940. LAPLAUD, M., AND CASSOU, R. ~qouveau Procede de Recolte du Sperme par Electrode Bipolalre Rectale Unique. Comp. rend. acad. agr. France, 31: 37. 1945. LAPLAUD, M., AND CASSOU, R. Sur la Production de Trois Veaux Normands, ~ 1' Aide de Sperme de Taureau Recueilli Electriquement, Dilu@, Conservd et Transport6. Comp. rend. acad. sci. Paris, 229: 629. 1949.
ELECTROEJACULATION IN DAIRY BULLS
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(9) LAPLAUD, •., ORTAVANT, •., AND THIBAULT, C. L'Electroejaculation chez le Taureau peut-elle Devenir une Methode Courante de Collecte de Sperme? Comp. rend. acad. agr. France, 34: 731. 1948. (10) LAUlCANS, :R., AND CLE,i~[ENT~ P. Nouvelles Electrodes pour l'Electroejaculation des Ovins et de Bovins. Co~ap. rend. acad. agr. France, 36: 450. 1950. (11) LIKAR, I., AND KAMHI, S. Elektro-ejakulacija kod Ovna. Veternaria (Sarajevo) 1: 506. Anin~al Breed. Abstr. 2l, No. 274. 1953. (12) LVTWAK-MANN, C., AND ROWSON, L. A. The Chemical Composition of the Pre-Sperm Fraction of Bull Ejaculate Obtained by Electrical Stimulation. J. Agr. Sci., 43: 9. 1953. (13) ~¢[ASCARENHAS,T:I., AND CJOMES, W. V. Contribuieao ao Estudo da Electroejaculacao em Bovines. Novo Tipo de Eletrodo Bipolar e Tecnica de sua Aplicaeao. Publ. inst. zootec. (Rio de Janeiro) No. 8: 1. 1950. (14) MATA, E. GARCIA, AND CA_ixTO, A. E. L'Applieazionc dell Elettroei~eulazione ed il Controllo Della Fecondith. (La Fecondazionc Avtificiale) Zootech. e Vet., 7: 38. 1952. (15))/[OOKE, C. R., AND GALL&GHEI~, T. F. SeminaLVesicle and Prostate Function as a TestisHormone Indicator; the Electric Ejaculation Test. A ~ . J. Anat., 45: 39. 1930. (16) O~TAVANT, l~., L&PLAUD, )/[., AIqD THIBAULT, C. Influence de l'Electro-Ejaeulation sur la Qualiti du Sperme chez le Belier. Comp. rend. acad. agr. France, 34: 733. 1948. (17) TEn,ILL, C. E. Comparison of Ram Semen Collection Obtained by Three Different Methods for Artificial Insemination. Proc. Am. Soc. Animal Prod. p. 201. 1940. (18) THIBAULT, C., L&PLAUD, M., AND ORTAVA.NT, R. L'Electro-Ejacul&tion chez le Taureau, Technique et R~sultats. Comp. rend acad. sci. Paris, 226: 2006. 1948. (19) THIBAULT, C. t'ersonal comn~unication, 1952.
AND
P. J. DZIUK, E. F. GRAHAM, AN]) W. E. PETERSEN
Departv~ent of Dairy Husband~'y University of Minnesota, St. 2aul The widespread use of artificial insemination has increased the value of each ejaculate of semen f r o m valuable sires. Since some males are either unwilling or unable to serve an artificial Vagina, their usefulness is limited. Electrical stimulation is one of several methods that have been used in an a t t e m p t to collect semen f r o m such animals. One of the first to record the obtaining of semen b y the use of electrical stimulus was Batelli (1), who collected f r o m guinea pigs. Moore and Gallagher (15) modified the technique and used it in an evaluation of the functioning of the accessory glands of the guinea pig. G u n n (5) was the first to make extensive use of electrical stimulus as a means of collecting semen f r o m rams. F u r t h e r work with this technique was done b y Bonadonna (2), L a m b e r t and 1VicKenzie (6), Terrill (17), and others. The stimulation was 60 cycle a l t e r n a t i n g current of about 30 volts applied b y means of a rectal probe and a l u m b a r electrode. L a p l a u d and Cassou (7) constructed a bipolar rectal probe which p e r m i t t e d stimulation without an external l u m b a r electrode. L a u r a n s and Clement (10), Likar and K a m h i (11), Mata and Cano (14), and O r t a v a n t et al. (16) made f u r t h e r use of the bipolar electrode in collecting semen f r o m rams. The bipolar probe was a d a p t e d b y Thibault et al. (18) so t h a t it could be used in the stimulation of bulls. His equipment consisted of a probe with 30 brass rings insulated f r o m each other by ebony. H e used 60 cycle a l t e r n a t i n g c u r r e n t which was varied f r o m zero to t h i r t y volts and back to zero every 3 to 5 seconds. Evidence is given on p i t , volume, density, and motility of the semen to indicate t h a t the semen samples collected were normal. Thibault (19) described i~l detail the electrical circuits, construction of the probe, and the technique used in r e s t r a i n i n g the bull and collecting the semen. L a p l a u d e t al. (9) f u r t h e r elaborated on the application of electrical stimulus as a method of obtaining semen f r o m bulls. L a p l a n d and Cassou (8) cite an example of a young bull that had yielded 161 ejaculates in an artificial insemination center before an accident which affected his h i n d q u a r t e r s p r e v e n t e d a n y a t t e m p t at service. A 6-ml. sample of semen was obtained b y bipolar electrical stimulation and used to inseminate two cows. A pair of f r a t e r n a l twins and a single calf were b o r n f r o m these matings. L u t w a k - M a n n and Rowson (12) made several chemical determinations of various fractions of bull semen collected b y electrical stimulus. They found t h a t the p r e - s p e r m fraction was not f r o m Received for publication February 1, 1954. 1University of Minnesota Scientific Journal Series Paper No. 3108, i~innesota Agricultural :Experiment Station. 1035
1036
e - J. D Z I U K E T AL
the accessory glands but p r o b a b l y f r o m the urethral glands. Excesses of this fluid did not affect motility or fructolysis. E g r o n (4) f o u n d the semen collected by means of artificial vagina did not differ greatly f r o m t h a t obtained b y electrical stimulus. The p t I was slightly higher and the density of s p e r m was lower in semen resulting f r o m electrical stimulation. Teasing the bull before collection increased the density. Briere (3) mentioned the applications of eleetroejaculation in his thesis. Mascarenhas and Gomes (13) were the first to report large numbers of collections by electrical stimulation in the bull. They used the multi-ringed bipolar probe to a p p l y both constant and impulse type of current. The impulse type of c u r r e n t is described in which the stimulation is passed f r o m one group of rings to another in r a p i d succession. This type of stimulation gave less fluid and the side reactions were more marked than with the constant type. Initial experiments were p e r f o r m e d by using a 15-v. current, b u t later work was done with a current of 5-10 v., which also proved satisfactory. A total of 48 separate collections were made f r o m seven bulls. Generally, the semen was less dense but of higher volume so that the total numbers of sperm were essentially the same as in semen collections made with an artificial vagina. E X P E R I M E N T A L PROCEDURE
Eighteen bulls were used as experimental animals in these trials. Holstein, Jersey, Guernsey, Brown Swiss, and Shorthorn breeds were represented and, except for a set of f r a t e r n a l Holstein twins, all were p u r e b r e d animals. Several different types of electrical stimuli were tried with v a r y i n g degrees of success. The o r d i n a r y inductorium stimulus of high voltage peaks and low a m p e r a g e was found to be highly stimulating to motor nerves, especially the sciatic, but was ineffective in causing either erection or ejaculation. The frequency of the stimulating current was varied f r o m 15 to 900 cycles per second b y means of a variable frequency oscillator. No advantage was found for either the higher or lower frequencies. Because of the expense and inconvenience of the equipment, s t a n d a r d 60-cycle current was used subsequently. A 30-v., 60-cycle alternating current which could be varied f r o m zero to 30 v. by means of a rheostat also was tried. Because of undesirable fluctuations in the voltage applied to the bull as the c u r r e n t load increased, the rheostat was discarded in favor of a variable t r a n s f o r m e r as a means of v a r y i n g the voltage level. An isolation step-down t r a n s f o r m e r was found to be necessary in isolating the circuit from the power source and to allow more gradual changes in voltage. A t t e m p t s were made to include resistors v a r y i n g from 25 to 200 ohms in series with the bull in the final circuit. This was done to reduce marked changes in the current flow through the bull as the resistance changed because of feces accumulation and poor contact. Some mechanism, p e r h a p s contraction of the rectum upon stimulation, causes the effective resistance of the bull to decrease as the a m p e r a g e flow increases. Therefore, the added series resistor prevented a sufficiently high amperage level to achieve either an erection or an ejaculation.
ELECTROEJACULATION IN DAIRY BULLS
1037
Several rectal probes of various dimensious and specifications were used in these trials. The early a t t e m p t s were made with only two electrodes. These were moved and placed b y insertion of the gloved a r m into the rectum. B y localizing the stimulation over the glands and o v e r the bulbo-cavernosus muscle, pronounced erection and a v e r y little fluid could be obtained; however, in no case was an ejaculatiou obtained. One probe 1 in. in diameter with seven rings 13A in. a p a r t was used with some success but did not give consistent results. A second probe, 1 in. in diameter with nine rings 11A in. apart, gave much more fluid, but ejaculation did not follow in all cases. A smaller probe, 3/~ in. in diameter with six rings 3A in. apart, gave v e r y little stimulation even with the resulting high amperage. A probe 2 in. in diameter with three longitudinal electrodes was used but was found to be less desirable t h a n the ringed probe of similar dimensions, because of more side reactions and more fluid secretion. The probe which seemed most satisfactory was 11/2 in. in diameter with six rings 13~ in. apart. This probe was made f r o m a section of 11/2 in. h e a v y steam hose 2 feet long. E a c h ring electrode was p a r t i a l l y inlaid into the rubber. The electrodes were made f r o m 2 turns of h e a v y solid core solder. A separate lead r a n inside the probe and was soldered to each electrode. Most of the results reported were obtained with this t y p e of probe and the electrical circuit shown in F i g u r e 1. II0/30-Volt transformer
~,,2-Amp fuse ~
\ \ \ \ " \ "--Ammeter ' ~_ ~
22"
>1
,oT¢ l
~Voltmeter Rectal probe ~-Varioble transformer
FIG. 1. Schematic diagram of stimulator. Collection of the semen was made b y means of a r u b b e r cone which was held open by a section of radiator hose 11/~ in. long, similar to t h a t used for the artificial vagina. The opening was protected fronl dirt b y a stiff r u b b e r sheet with several intersecting slits cut in it. The semen was collected in a test tube which was s u r r o u n d e d b y a larger test tube to serve as a w a r m w a t e r jacket to minimize cold shock. I t is necessary to be able to see the semen at collection to aid in deciding on the n u m b e r and intensity of subsequent stimulations. The technique used became f a i r l y well established as the trials progressed. The first step is administration of an enema of about two quarts of a w a r m 5% solution of sodium chloride. I t is i m p o r t a n t to give an adequate enema to remove as much of the fecal material as possible in order to minimize its insulation of the electrodes. The p r e p u t i a l area should be clipped and brushed to reduce the amount of d i r t and foreign material which otherwise m a y get in the semen. The probe is lightly lubricated with some noninsulating material a n d introduced into the rectum. The probe is inserted f o r v a r y i n g distances,
1038
P. $. D Z I U K ET AL
d e p e n d i n g on the size of the bull, b u t g e n e r a l l y no more t h a n 12 in. The best results, especially as f a r as e r e c t i o n is concerned, were g e n e r a l l y o b t a i n e d w h e n the p r o b e was m a i n t a i n e d on the v e n t r a l side of the r e c t u m . The voltage was g r a d u a l l y i n c r e a s e d to a b o u t 3-5 v. a n d t h e n back to zero e v e r y 5-10 seconds. The s u b s e q u e n t s t i m u l a t i o n s go p r o g r e s s i v e l y h i g h e r so t h a t at a b o u t the fifth s t i m u l a t i o n the m a x i m u m of 10-15 volts is reached. There is m u c h v a r i a t i o n b e t w e e n bulls, a n d f r o m one time to a n o t h e r w i t h the same bull, so t h a t e x p e r i e n c e best g o v e r n s the r a p i d i t y of increase. E r e c t i o n a n d fluid secretion u s u a l l y s t a r t at a b o u t the 5:v. level, b u t e j a c u l a t i o n u s u a l l y occurs at the 10-15 v. m a x i m u m , a t which time a b o u t 0.5 to 1.0 a m p e r e of c u r r e n t is flowing. Too m a n y s t i m u l a t i o n s a t a s u b - e j a c u l a t o r y level y i e l d m u c h fluid a n d o f t e n make i t difficult to o b t a i n a n e j a c u l a t i o n at a n y level. Too r a p i d a n increase m a y lead to a p r e - e r e c t i o n e j a c u l a t i o n a n d c o n t a m i n a t i o n of the semen w i t h dirt, since the p e n i s is n o t free of the sheath. A t the time of s t i m u l a t i o n m a n y m o t o r n e r v e s are also s t i m u l a t e d , a n d g e n e r a l t e t a n y of the h i n d legs results. T h e r e is a g e n e r a l e x t e n s i o n of the h i n d legs a n d a t e n d e n c y to p u s h f o r w a r d . However, i n n o ease has a b u l l been observed to establish u n f a v o r a b l e associations w i t h e i t h e r the collection stall or the m e t h o d of collection. I t was f o u n d t h a t a m i n i m u m of r e s t r a i n t was n e c e s s a r y f o r best collection, the o n l y r e q u i r e m e n t b e i n g a s t u r d y s t a n c h i o n or stock to r e s t r a i n the neck a n d to allow the b u l l to p u s h f o r w a r d , Good footing, such as e a r t h or gravel, was needed, since the b u l l s s l i p p e d on concrete or wooden floors. TABLE 1 Collection data
Bull No.
No. of successful electroejaculations
:No. of attempts
Total volume of fractions containing sperm
Average volume
1 2 3 4 5
29 2 3 0 1
30 3 4 3 1
(ml.) 327 33 30 0 10
(ml.) 11 i6 10 0 10
6
6
6
74
12
7 8 9 10 11 12 13 14 15 16 17 18 19
9 13 9 12 12 2 3 1 2 2 3 1 1
10 13 10 12 12 2 3 1 2 2 3 1 1
93 189 216 95 77 32 36 9 33 39 63 8 11
10 14 24 8 6 16 12 9 16 19 21 8 11
Total
111
119
1,375
ELECTROEJACULATION IN DAIRY BULLS
1039
RESULTS AND DISCUSSION
One h u n d r e d eleven s e p a r a t e collections were m a d e f r o m 18 d i f f e r e n t bulls. S e m e n was n o t o b t a i n e d in e i g h t a t t e m p t s . Of t h e 19 b u l l s used, o n l y one f a i l e d to y i e l d a n y semen. T h r e e s e p a r a t e a t t e m p t s w e r e m a d e w i t h t h i s bull, b u t d e a t h d u e to p n e u m o n i a p r e v e n t e d f u r t h e r a t t e m p t s . T h e v o l u m e of s e m e n o f t h e 111 s u c c e s s f u l collections a v e r a g e d 12.3 ml. w i t h a r a n g e of f r o m 1 to 35 ml. ( T a b l e 1). The d e n s i t y r a n g e d f r o m v e r y low to 1.27 b i l l i o n p e r m i l l i l i t e r i n a l m o s t a n i n v e r s e r e l a t i o n s h i p to volume. T h e e x t e n t of s t i m u l a t i o n a n d d e g r e e of s e p a r a t i o n a r e t h e g o v e r n i n g f a c t o r s of t h e d e n s i t y - v o l u m e r e l a t i o n s h i p . U s u a l l y , no a t t e m p t was m a d e to f r a c t i o n a t e t h e fluid p o r t i o n s f r o m t h e m o r e dense semen, since it was b e l i e v e d t h a t the excess fluid was n o t so d e t r i m e n t a l to t h e s e m e n as was t h e n e c e s s a r y i n t e r r u p t i o n i n t h e sequence of collection w h e n t h e f r a c t i o n s were s e p a r a t e d . O f t h e u n s e l e c t e d s a m p l e s in w h i c h t h e t o t a l n u m b e r of s p e r m was d e t e r m i n e d , t h e r e was no a p p r e c i a b l e difference i n t o t a l n u m b e r s of s p e r m b e t w e e n s a m p l e s collected b y m e a n s of a r t i f i c i a l v a g i n a a n d those collected b y e l e c t r i c a l s t i m u l a t i o n . A l t h o u g h t h e r e w e r e i n d i v i d u a l b u l l differences, t h e first few s t i n m l a t i o n s u s u a l l y y i e l d e d some fluid, f o l l o w e d b y t h e m o r e c o n c e n t r a t e d semen. F u r t h e r s t i m u l a t i o n y i e l d e d m o r e d i l u t e s e m e n a n d f i n a l l y c l e a r fluid. One n o r m a l b u l l was collected f r o m b y e l e c t r i c a l s t i m u l u s a b o u t once w e e k l y f o r a t o t a l of 30 c o l l e c t i o n s ( T a b l e 2). H e f o r m e d no u n TABLE 2 Evaluation of unselected semen samples collected by electroejaculation from buU No. 2
Collection :No.
Total volume
Total sperm :No. in billions
(ml.) 1 2 3 4 5 6 7 8 9 10 a 11 12 13 14
5 9 18 31 13 5 4 15 12 10 7 20 5 11
Motility rate (%)
5.0 5.7 7.0 15.5 16.4 7.6 1.2 10.3 9.6 10.5 6.4 21.0 4.0 13.4
85 85 95 85 85 85 80 85 75 95 80 85 50 80
a Collected by artificial vagina 5 minutes after collection :No. 9. f a v o r a b l e a s s o c i a t i o n w i t h the t r e a t m e n t a n d was n o t affected in a n y o t h e r w a y . H e did, on s e v e r a l occasions, serve a n a r t i f i c i a l v a g i n a w i t h i n a f e w m i n u t e s a f t e r collection b y e l e c t r o e j a c u l a t i o n a n d p r o d u c e d a n o r m a l e j a c u l a t e . N i n e of t h e b u l l s i n a r t i f i c i a l i n s e m i n a t i o n s t u d s were i m p o s s i b l e to collect f r o m w i t h t h e a r t i f i c i a l v a g i n a . F o r t y - s e v e n collections w e r e m a d e f r o m these bulls. T w e n t y - s e v e n of these collections were d i l u t e d a t u s u a l r a t e s a n d u s e d in t h e field d u r i n g t h e m o n t h s of M a r c h , A p r i l , M a y , a n d N o v e m b e r , 1953. R e s u l t s
P . J. D Z I U K
1040
ET AL
TABLE 3 Conception data on bulls which were electroejaculated
Bull
A
B
C
D
Number of first services 60-90-day nonreturns ( % ) Previous year over-all ( % ) 60-90 day nonreturns
28 68.0 67.0
2,012 60.4 61.9
138 44.9 55.0
89 57.3 61.5
o f t h i s t r i a l a r e s h o w n i n T a b l e 3. T h e c o n c e p t i o n d a t a f o r t h e r e m a i n i n g c o l l e c t i o n s is n o t a v a i l a b l e at t h e t i m e of w r i t i n g .
10
SUMMARY AND CONCLUSIONS E v i d e n c e h a s b e e n p r e s e n t e d w h i c h i n d i c a t e s t h a t i t is p o s s i b l e b y e l e c t r i c a l s t i n m l u s to c o l l e c t s e m e n f r o m s o m e b u l l s w h i c h a r e e i t h e r u n a b l e o r u n w i l l i n g to s e r v e a n a r t i f i c i a l v a g i n a . T h e r e h a v e b e e n n o a d v e r s e effects o r u n f a v o r a b l e a s s o c i a t i o n s i n b u l l s c o l l e c t e d f r o m b y e l e c t r o e j a c u l a t i o n . T h e s e m e n so o b t a i n e d w a s g e n e r a l l y of g r e a t e r v o l u m e b u t of l o w e r d e n s i t y . H o w e v e r , t h e t o t a l n u m b e r s of s p e r m a t o z o a w e r e c o m p a r a b l e to a n e j a c u l a t e o b t a i n e d b y a n a r t i f i c i a l vagina. E l e c t r o e j a c u l a t i o n m a y s e r v e as a v a l u a b l e a d j u n c t to t h e a r t i f i c i a l v a g i n a as a m e a n s of c o l l e c t i n g s e m e n i n a r t i f i c i a l i n s e m i n a t i o n c e n t e r s , e s p e c i a l l y f o r b u l l s w h i c h a r e d i f f i c u l t to c o l l e c t s e m e n f r o m b y m e a n s of t h e a r t i f i c i a l v a g i n a . ACKNOWLEDGMENT The authors wish to acknowledge the cooperation of John Dysart of Land O'Lakes Herd Improvement Division and Wallace Miller of Minnesota Valley Breeders' Association in making conception data available for this study. REFERENCES
(i) BATELLI, (2) (3) (4) (5)
(6)
(7) (8)
F.
Une
Methode
pour
obtenir
l ~ Emission
Complete
du Liquide
des Vesicules
Seminales chez le Cobaye. Soc. de Physique et d' Hist. nat. de Gcneve, Co~np. Rend., 39: 73. 1922. BONADONNA,T. I1 Metodo dell' Elettroeiaculazione negli Ovini e nei Caprini. La Fccondazione Artificiale, 1: 54. 1938. (Supp. to Clin. Vet., 61. 1938.) BRIEI~E, J. Contribution a l'Etude des M@thodes de Collecte du Sperme chez le Taureau. Thesis 1-43 Alfort. 1950. (Abs.) Vet. Bull. 319'~'. 1952. EGR0~I, .... Sur quelques R~sultats Pratiques de 1' Electro-Ejaculation. Elevage ct Inse~n. No. 13: 19. 1952. Ani~na! Breed. Abstr., 21, No. 218. 1953. GVNN, ]~. M. C. Artificial Production of Seminal Ejaculation and the Characters of the Spermatozoa Contained Therein. Council for Scientific and Industrial Research, Bull. 94. 1936. LAMBEI~T~W. V., AND MCKENZIE, F. F. Artificial Insemination in Livestock Breeding. USDA Circular 567. 1940. LAPLAUD, M., AND CASSOU, R. ~qouveau Procede de Recolte du Sperme par Electrode Bipolalre Rectale Unique. Comp. rend. acad. agr. France, 31: 37. 1945. LAPLAUD, M., AND CASSOU, R. Sur la Production de Trois Veaux Normands, ~ 1' Aide de Sperme de Taureau Recueilli Electriquement, Dilu@, Conservd et Transport6. Comp. rend. acad. sci. Paris, 229: 629. 1949.
ELECTROEJACULATION IN DAIRY BULLS
1041
(9) LAPLAUD, •., ORTAVANT, •., AND THIBAULT, C. L'Electroejaculation chez le Taureau peut-elle Devenir une Methode Courante de Collecte de Sperme? Comp. rend. acad. agr. France, 34: 731. 1948. (10) LAUlCANS, :R., AND CLE,i~[ENT~ P. Nouvelles Electrodes pour l'Electroejaculation des Ovins et de Bovins. Co~ap. rend. acad. agr. France, 36: 450. 1950. (11) LIKAR, I., AND KAMHI, S. Elektro-ejakulacija kod Ovna. Veternaria (Sarajevo) 1: 506. Anin~al Breed. Abstr. 2l, No. 274. 1953. (12) LVTWAK-MANN, C., AND ROWSON, L. A. The Chemical Composition of the Pre-Sperm Fraction of Bull Ejaculate Obtained by Electrical Stimulation. J. Agr. Sci., 43: 9. 1953. (13) ~¢[ASCARENHAS,T:I., AND CJOMES, W. V. Contribuieao ao Estudo da Electroejaculacao em Bovines. Novo Tipo de Eletrodo Bipolar e Tecnica de sua Aplicaeao. Publ. inst. zootec. (Rio de Janeiro) No. 8: 1. 1950. (14) MATA, E. GARCIA, AND CA_ixTO, A. E. L'Applieazionc dell Elettroei~eulazione ed il Controllo Della Fecondith. (La Fecondazionc Avtificiale) Zootech. e Vet., 7: 38. 1952. (15))/[OOKE, C. R., AND GALL&GHEI~, T. F. SeminaLVesicle and Prostate Function as a TestisHormone Indicator; the Electric Ejaculation Test. A ~ . J. Anat., 45: 39. 1930. (16) O~TAVANT, l~., L&PLAUD, )/[., AIqD THIBAULT, C. Influence de l'Electro-Ejaeulation sur la Qualiti du Sperme chez le Belier. Comp. rend. acad. agr. France, 34: 733. 1948. (17) TEn,ILL, C. E. Comparison of Ram Semen Collection Obtained by Three Different Methods for Artificial Insemination. Proc. Am. Soc. Animal Prod. p. 201. 1940. (18) THIBAULT, C., L&PLAUD, M., AND ORTAVA.NT, R. L'Electro-Ejacul&tion chez le Taureau, Technique et R~sultats. Comp. rend acad. sci. Paris, 226: 2006. 1948. (19) THIBAULT, C. t'ersonal comn~unication, 1952.