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Sperm concentration in different segments of the goat epididymis
THERIOGENOLOCY
SPERM
CONCENTRATION
IN DIFFERENT EPIDTDYMIS
SEGMENTS
nF THE
G3AT
S.K.Jindal & Climatology Division OP Physiology Tndian Vetti~inary Research Institute Izatnag.-lr - 243 122 INDIA Received
for
publication: Accepted:
7, 1984 August 20, 1984 March
ABSTR.4C'T Spermatocrit values of fluid coLlected by micropuncture from the caput, corpus and cauda of goat epididymides as well as rete testis fluid and testicular fluid were determined for eight animals. The values indicatsd that 98.02% of fLui;l is absorbed in the epididfmis and that the cauda epididymis contains the lowest amount of fluid, significantly different from the rete testis, corpl~s and caput. Keywords
: Sperm crit,
concentration, Goat.
Epididymis,
Spermnto-
INTROD3CTION Spermatozoa mature along the epididymal ducts befo-e they are capabls of fertilization. When mat ore spermatozoa acquire motility, movemeut of cytoplasmic droplet and other morphological changes (1,Z). The epididymis influences the spermatozoa via the sperm cell environment i.e. the epididymal plasma. Sperm concentration in different parts of the epididymis has been used to calcrllate the c‘hanges in volu.ne of -the epididymn' contents from segment to segment. This technique is valid only when we assume that the absolute number of spermatozoa rema'ns constant during transit through the epididymis. Several studies have there are reports to the coilprov*?n this, although traty. Sperm concentration in different parts of
--_-------_____ Acknowledgement
NOVEMBER
: The
author is grateful to Dr. N.K. Bhattacharyya for providing the necessary facilities for carrying out this work.
1984 VOL. 22 NO. 5
545
THERIOGENOLOGY
epididymis has been reported for the bull (3), boar bull (5) and rat (6-p). This study was (4), buffalo designed to report this information for the goat. MITERIALS
AND
MZ'TH3DS
The methods follow-d were essentially similar to those of Sharma et al (5) and Crabo (4) with minor modifications. FS‘eszs fro.n adult goaks were collected immediately after slaughter and carried to the laboratory under ice in a vacuum flask. Laboratory proce+ occurred within a? hour after slaughter. The sim epididymis xas removed from the testis and divided into cap11t ) corpus aid cauda on the basis of external morphology. The f1ui.d collection sites from the caput, co.:pus and cauda coresponded with regions D, E and F, respectively, as used by Crabo (4) for the bull. Capillary tubes with an outside diameter of 1 mm and inside diameter of 0.72 mm were a:jed to collect tha? fluid after the seminifero.us tubules were cut. and squeezed. The tips were sealed on one sicle by heating the end on a spirit lamp flame, care not to affect the contents inside the taking The tubes were marked to identify the sample tube. and centrifuged in a Remi Microhematocrit centrifuge model R-18 (manufactured by Remi India, New Delhi, The spermatocrit India) for 3 l/2 min at full speed. of the total value was expressed as the percentage No correction was salnple. voLume of the aspirdted trapping. ma?e for plasma Blood was collected from the veins 0; the pampiniform plexus into heparinized capillary tubes. The rete testis fluid was collected by squeezing the testis immediately after the epididymis jzlasremoved and aspirating the clear fluid, uncontaminated by The testithat came out of the rete testis. blood, cular fluid was collected by aspirating the fluid after cutting open the testis. The milky white fluid is supposedly a mixture of testicular lymph tubule fluid. and seminiferous RESULTS
AND
DlSCUSSION
The spermatocrit values in the testicular fluid, rete testis fluid, caput, corpus and cauda epididymal fluid and homatocrit values are shown in Table 1. The
546
NOVEMBER
1984 VOL. 22 NO. 5
THERIOGENOLOGY
values for rete testis fluid, corpus and testicular fluid are reported for only a few animals because we had earlier planned to study onl'y the spermatocrit value of caput and cauda parts. Th,z spcrmatocrit value of the caput epididymaL fluid is 21.20, mucn higher than the rete testis fluid value of 1.65. spermntocrit value of the cauda epididymal The fluid was 45.89 which was aLso higner tha? the corpus and caput epididymis fluid values, 22.93 and 21.20 respectively.
Table
Spermatocrit in goat epididymis cular blood.
1.
Rete testis fluid
Testis fluid
of the of testi:
Corpus
Cauda
Blood
21.20 (16)
22.93 (7)
45.89 (15)
20.65 (113)
3.19
2.03
2.64
0.57
CapuI
1.65 (9)
42.01 (7)
various poj-tions and haematocrit
SEM 3.48
't'
0.60
value
*P
4.38*
in
0. 33NS
NS = Not significant. Inumber parenthesis indicat?
5.34*
of
observalions.
spermatozoa are bathed in the epidldymis, ln the fluid OI- epid:dymal plasma that fills the lumen of This fluid is deri~Jed from the rete testis the duct. fluid but is also modified by the secretions and absorption by th,? epidldymal epithelium. Our results are consistent with the findings of earlier workers (6, 10) that substantial resorption of fluid occurs in the proximal reg.ion of the epididymis. BecaTlse of this fluid resorption, spermatozoa is concentrated 10. This increases the chances that fold or more. spermatozoa will interact with the m,aturation factors and also brings such facto-s present in either the rete testis fluid or secreted by the epididymis, to critical concentrations. Absorption in the proximal capu-t starts the maturation which culminates in the distal corpus and/or proximal cauda, in the goat.
NOVEMBER
1984VOL.22N0.5
547
THERIOGENOLOGY
As reported earlier (2), progressive motility was found only in the distal corpus with a gradation in the increase of spermatozoa sho,nring progressi-se motility starting from distal part of the corpus and culminating at the distal lobe of the cauda. Jessee and Howards (11) object to using spermatocrit values to calculate fluid absorption, because the spermatocrit is affected by the changing physio-chemical properties of the sperm and fluid and the amount of fluid trapped in the sperm phase is not linearly related to the concentra'ion of sperm. However the use of otaer techniques, like counting the sperm concentration directly, by these authors has not changed the gross conclusions previously obtained in the rat or other species. by spermatocrit values value in the goat cauda The mean spermatocrit (45.89) is lower than the values reported for caquda and guinea pig, epididymal plasma of rat, hamster 52.86, 43.45 and 49.61 respectively (12) and for a It does, similar region in elephant, 50.37 (13). correlate well with the valua of the bull, however, it is much higher than that 47.6 (4), although The reareported for the buffalo bull, 56.13 (5). Crabo (4) sons for this discrepancy are not obvious. has also reported a much higher value of 52.3 for The rete testis fluid spermatocrit in distal cap?Jt. corresponds well with value for goa? spermatocrit the value of 1.2 reported for the elephant (13). Crabo (4) reported a value of 2.8 for fluid collected from similar region in the bull. The volume of the fluid prorluced ever) day by the. ram t.cst-is is approximotcl y 100 times tba 1 whi c11 in a sexuaily active anim,al the epididymis leaves This means that epididymal fluid resorp(1i--16). We estimate from the present data tion does occur. that 94.7646 of the fluitl in goats was absorbed dut'ing the passage from the rete testis to the caput epididymis and while there .was a small insignificant decrease of 0.59$ from caput to car-pus, there was a further absorption ol‘ 3.6'746 from corpl~s to cauda as 0,' epididymnl shown by the talc-ulated relatiqre volume content for the v-arious segments of goat epid:dymis. plasma volume in So in all, 98.02% of the epididymal i- the epididymis. Wong rete testis .&as absorbed (17) found that the fluid absorption is and Yeunq but that it is secoilaary to the energy dependent,
548
NOVEMBER
1984VOL.22NO.5
THERIOGENOLOGY active transport ol-‘sodium. The sperm dehydration may also help form disulphide (S-S) bridges between the thiol(SH-) containing cysteine residites of the sperm histones. Kvist (13) states that the formation oi" S-S bridges apparently commences in the epididymal caput and no-mally continues dllring the sperm transport throllsh the remainder of the epidia definite proof that f‘luid absorpdymis. However, tion is a necessary prerequisite for matur.jtion is still missing. REFERENCES 1.
transporti and fate of Bedford, J.M . Maturation, spermatozoa in the epididymis. In: Hamilton, Handbook of PhysioR.O., D.W. and GreeD, (ed) logy, Vol. V Section 7, Ma12 Reprodllective System, Waverly Press, Baltimore, 1975, pp. 30’3-318.
2.
Jindal, S.K. and Zanda. J.N. Maturat ion chan,:?s of goat spe.?matozoa during transit throtlgh thl(I 180). epididymis. Andrologia, -12~328-331
3.
B. Distribxtion oZ Crabo, B. and Gustaffsoil, sodium and potassium and its relation to sperm concentration in the epididymal pLasma of the hall. J. Reprod. Fertil. 2:337-345 (1*264).
4.
Crabo, B. Studizs on the compns-;tion ma1 co;ltent in bulls and boars. Acta Stand. Suppl. J:l-94 (1965).
5.
Sharma, A.K., Chaudhary, P.S. and Gupta, R.C. Sperm concentration in different paTts of the epididymis of buffalo bulls by micropuncture technique, AndroLogia (1079). -11:153-156
6.
Levine, N. and :4arsil, D.J. Micropuncture studies 0: the electrochemical aspects of the fluid and electrolyte transport of indivldllal seminiferous tubules, the epidldymis and vas deferens of the rat. J. Physiol. 213:557-570 (1971).
7.
Howards, S.S., Johnson, A. and .Jessee, S. Micropuncture and microanalytic studies o; rat testis and epididymis. Fertil. Steril. &:13-13 (1975).
NOVEMBER
1984 VOL. 22 NO. 5
ol' epididyVet.
the
549
THERIOGENOLOGY
8.
and Setchell, Hinton, B.T. glycerylphosphorylcholine, inorganic phosphate in the testis and epidldymis. J. 406
Concentration of B.P. phosphocholine and free lumina fluid of the rat Repr0.d. Fertil. 58:401-
(1930).
9.
Setchell, B.P. and Hinton, the spermatozoa of changes luminal fluids as it passes Progress in Reproductive In: Physiology of the Epididymis, pp. 58-66.
The effect on B.T. in the composition Of along the epididymis. Biology, Vol. 8. Karger, Basel, 1981,
10.
Turner, T.T., Hattman. potassium vivo sodium, the rat epididymis.
and Howsrds, S.S. sperm concentration Steril. -28:191-194
P.K. and Fertil.
In in
(1377). 11.
Jessee, S.J. and Howards, S.S. potassium and sodium concentration fluid of the hamster epididymis.
626-631
(1976).
A
surve" of sperm in the tubular Biol. Reprod. -15:
12.
Back, D.J. composition epididymis Experientia
and Shenton, J.C. A comparison of epididymal plasma from the of the rat, hamster and guinea (1975). -31:464-465
13.
Jones, R.C. spermatozoa elephant.
Luminal composition and maturation in the genital ducts of the African J. Repr0.d. Fertil. (1980). -60:87-93
1 G.
Setchell, B.P., Scott, T.W., Voglmayr, J.K. and Waites, G.M.H. Characteristics of testicular spermatozoa and the fluid which transports them into the epidldymis. Biol. Reprod. Suppl. 1: 40-60 (1969).
15.
Voglmayr, J.K., Waites, G.M.H. and Setchell, B.P. Studies on spermatozoa collected directly from the testis of the conscious rams. Nature 210:
861-863
16.
of the cauda pig.
of
(1964).
Voglmayr, J.K., Scott, T.W., qetchell, B.P. and Waites, G.M.H. Metabolism of testicular spermatozoa and characteristics of testicular fluid collscted from conscious rams. J. Reprod. Fertil.
-14:87-99
(1947).
NOVEMBER
1984 VOL. 22 NO. 5
THERIOCENOLOGY
17.
Wow,
18.
Kvist, IJ. Sperm nuclear chro‘natin decondensation ability. Acta. Physiol. Stand. Suppl. 436 : 1-24 (1980).
P.Y.D. tion in the epididymis.
NOVEMBER
Fluid reabsorpand Yeung, C.H. isolated duct of the rat cauda J. Reprod. Fertil. (1377). -43:77-81
1984 VOL. 22 NO. 5
551
SPERM
CONCENTRATION
IN DIFFERENT EPIDTDYMIS
SEGMENTS
nF THE
G3AT
S.K.Jindal & Climatology Division OP Physiology Tndian Vetti~inary Research Institute Izatnag.-lr - 243 122 INDIA Received
for
publication: Accepted:
7, 1984 August 20, 1984 March
ABSTR.4C'T Spermatocrit values of fluid coLlected by micropuncture from the caput, corpus and cauda of goat epididymides as well as rete testis fluid and testicular fluid were determined for eight animals. The values indicatsd that 98.02% of fLui;l is absorbed in the epididfmis and that the cauda epididymis contains the lowest amount of fluid, significantly different from the rete testis, corpl~s and caput. Keywords
: Sperm crit,
concentration, Goat.
Epididymis,
Spermnto-
INTROD3CTION Spermatozoa mature along the epididymal ducts befo-e they are capabls of fertilization. When mat ore spermatozoa acquire motility, movemeut of cytoplasmic droplet and other morphological changes (1,Z). The epididymis influences the spermatozoa via the sperm cell environment i.e. the epididymal plasma. Sperm concentration in different parts of the epididymis has been used to calcrllate the c‘hanges in volu.ne of -the epididymn' contents from segment to segment. This technique is valid only when we assume that the absolute number of spermatozoa rema'ns constant during transit through the epididymis. Several studies have there are reports to the coilprov*?n this, although traty. Sperm concentration in different parts of
--_-------_____ Acknowledgement
NOVEMBER
: The
author is grateful to Dr. N.K. Bhattacharyya for providing the necessary facilities for carrying out this work.
1984 VOL. 22 NO. 5
545
THERIOGENOLOGY
epididymis has been reported for the bull (3), boar bull (5) and rat (6-p). This study was (4), buffalo designed to report this information for the goat. MITERIALS
AND
MZ'TH3DS
The methods follow-d were essentially similar to those of Sharma et al (5) and Crabo (4) with minor modifications. FS‘eszs fro.n adult goaks were collected immediately after slaughter and carried to the laboratory under ice in a vacuum flask. Laboratory proce+ occurred within a? hour after slaughter. The sim epididymis xas removed from the testis and divided into cap11t ) corpus aid cauda on the basis of external morphology. The f1ui.d collection sites from the caput, co.:pus and cauda coresponded with regions D, E and F, respectively, as used by Crabo (4) for the bull. Capillary tubes with an outside diameter of 1 mm and inside diameter of 0.72 mm were a:jed to collect tha? fluid after the seminifero.us tubules were cut. and squeezed. The tips were sealed on one sicle by heating the end on a spirit lamp flame, care not to affect the contents inside the taking The tubes were marked to identify the sample tube. and centrifuged in a Remi Microhematocrit centrifuge model R-18 (manufactured by Remi India, New Delhi, The spermatocrit India) for 3 l/2 min at full speed. of the total value was expressed as the percentage No correction was salnple. voLume of the aspirdted trapping. ma?e for plasma Blood was collected from the veins 0; the pampiniform plexus into heparinized capillary tubes. The rete testis fluid was collected by squeezing the testis immediately after the epididymis jzlasremoved and aspirating the clear fluid, uncontaminated by The testithat came out of the rete testis. blood, cular fluid was collected by aspirating the fluid after cutting open the testis. The milky white fluid is supposedly a mixture of testicular lymph tubule fluid. and seminiferous RESULTS
AND
DlSCUSSION
The spermatocrit values in the testicular fluid, rete testis fluid, caput, corpus and cauda epididymal fluid and homatocrit values are shown in Table 1. The
546
NOVEMBER
1984 VOL. 22 NO. 5
THERIOGENOLOGY
values for rete testis fluid, corpus and testicular fluid are reported for only a few animals because we had earlier planned to study onl'y the spermatocrit value of caput and cauda parts. Th,z spcrmatocrit value of the caput epididymaL fluid is 21.20, mucn higher than the rete testis fluid value of 1.65. spermntocrit value of the cauda epididymal The fluid was 45.89 which was aLso higner tha? the corpus and caput epididymis fluid values, 22.93 and 21.20 respectively.
Table
Spermatocrit in goat epididymis cular blood.
1.
Rete testis fluid
Testis fluid
of the of testi:
Corpus
Cauda
Blood
21.20 (16)
22.93 (7)
45.89 (15)
20.65 (113)
3.19
2.03
2.64
0.57
CapuI
1.65 (9)
42.01 (7)
various poj-tions and haematocrit
SEM 3.48
't'
0.60
value
*P
4.38*
in
0. 33NS
NS = Not significant. Inumber parenthesis indicat?
5.34*
of
observalions.
spermatozoa are bathed in the epidldymis, ln the fluid OI- epid:dymal plasma that fills the lumen of This fluid is deri~Jed from the rete testis the duct. fluid but is also modified by the secretions and absorption by th,? epidldymal epithelium. Our results are consistent with the findings of earlier workers (6, 10) that substantial resorption of fluid occurs in the proximal reg.ion of the epididymis. BecaTlse of this fluid resorption, spermatozoa is concentrated 10. This increases the chances that fold or more. spermatozoa will interact with the m,aturation factors and also brings such facto-s present in either the rete testis fluid or secreted by the epididymis, to critical concentrations. Absorption in the proximal capu-t starts the maturation which culminates in the distal corpus and/or proximal cauda, in the goat.
NOVEMBER
1984VOL.22N0.5
547
THERIOGENOLOGY
As reported earlier (2), progressive motility was found only in the distal corpus with a gradation in the increase of spermatozoa sho,nring progressi-se motility starting from distal part of the corpus and culminating at the distal lobe of the cauda. Jessee and Howards (11) object to using spermatocrit values to calculate fluid absorption, because the spermatocrit is affected by the changing physio-chemical properties of the sperm and fluid and the amount of fluid trapped in the sperm phase is not linearly related to the concentra'ion of sperm. However the use of otaer techniques, like counting the sperm concentration directly, by these authors has not changed the gross conclusions previously obtained in the rat or other species. by spermatocrit values value in the goat cauda The mean spermatocrit (45.89) is lower than the values reported for caquda and guinea pig, epididymal plasma of rat, hamster 52.86, 43.45 and 49.61 respectively (12) and for a It does, similar region in elephant, 50.37 (13). correlate well with the valua of the bull, however, it is much higher than that 47.6 (4), although The reareported for the buffalo bull, 56.13 (5). Crabo (4) sons for this discrepancy are not obvious. has also reported a much higher value of 52.3 for The rete testis fluid spermatocrit in distal cap?Jt. corresponds well with value for goa? spermatocrit the value of 1.2 reported for the elephant (13). Crabo (4) reported a value of 2.8 for fluid collected from similar region in the bull. The volume of the fluid prorluced ever) day by the. ram t.cst-is is approximotcl y 100 times tba 1 whi c11 in a sexuaily active anim,al the epididymis leaves This means that epididymal fluid resorp(1i--16). We estimate from the present data tion does occur. that 94.7646 of the fluitl in goats was absorbed dut'ing the passage from the rete testis to the caput epididymis and while there .was a small insignificant decrease of 0.59$ from caput to car-pus, there was a further absorption ol‘ 3.6'746 from corpl~s to cauda as 0,' epididymnl shown by the talc-ulated relatiqre volume content for the v-arious segments of goat epid:dymis. plasma volume in So in all, 98.02% of the epididymal i- the epididymis. Wong rete testis .&as absorbed (17) found that the fluid absorption is and Yeunq but that it is secoilaary to the energy dependent,
548
NOVEMBER
1984VOL.22NO.5
THERIOGENOLOGY active transport ol-‘sodium. The sperm dehydration may also help form disulphide (S-S) bridges between the thiol(SH-) containing cysteine residites of the sperm histones. Kvist (13) states that the formation oi" S-S bridges apparently commences in the epididymal caput and no-mally continues dllring the sperm transport throllsh the remainder of the epidia definite proof that f‘luid absorpdymis. However, tion is a necessary prerequisite for matur.jtion is still missing. REFERENCES 1.
transporti and fate of Bedford, J.M . Maturation, spermatozoa in the epididymis. In: Hamilton, Handbook of PhysioR.O., D.W. and GreeD, (ed) logy, Vol. V Section 7, Ma12 Reprodllective System, Waverly Press, Baltimore, 1975, pp. 30’3-318.
2.
Jindal, S.K. and Zanda. J.N. Maturat ion chan,:?s of goat spe.?matozoa during transit throtlgh thl(I 180). epididymis. Andrologia, -12~328-331
3.
B. Distribxtion oZ Crabo, B. and Gustaffsoil, sodium and potassium and its relation to sperm concentration in the epididymal pLasma of the hall. J. Reprod. Fertil. 2:337-345 (1*264).
4.
Crabo, B. Studizs on the compns-;tion ma1 co;ltent in bulls and boars. Acta Stand. Suppl. J:l-94 (1965).
5.
Sharma, A.K., Chaudhary, P.S. and Gupta, R.C. Sperm concentration in different paTts of the epididymis of buffalo bulls by micropuncture technique, AndroLogia (1079). -11:153-156
6.
Levine, N. and :4arsil, D.J. Micropuncture studies 0: the electrochemical aspects of the fluid and electrolyte transport of indivldllal seminiferous tubules, the epidldymis and vas deferens of the rat. J. Physiol. 213:557-570 (1971).
7.
Howards, S.S., Johnson, A. and .Jessee, S. Micropuncture and microanalytic studies o; rat testis and epididymis. Fertil. Steril. &:13-13 (1975).
NOVEMBER
1984 VOL. 22 NO. 5
ol' epididyVet.
the
549
THERIOGENOLOGY
8.
and Setchell, Hinton, B.T. glycerylphosphorylcholine, inorganic phosphate in the testis and epidldymis. J. 406
Concentration of B.P. phosphocholine and free lumina fluid of the rat Repr0.d. Fertil. 58:401-
(1930).
9.
Setchell, B.P. and Hinton, the spermatozoa of changes luminal fluids as it passes Progress in Reproductive In: Physiology of the Epididymis, pp. 58-66.
The effect on B.T. in the composition Of along the epididymis. Biology, Vol. 8. Karger, Basel, 1981,
10.
Turner, T.T., Hattman. potassium vivo sodium, the rat epididymis.
and Howsrds, S.S. sperm concentration Steril. -28:191-194
P.K. and Fertil.
In in
(1377). 11.
Jessee, S.J. and Howards, S.S. potassium and sodium concentration fluid of the hamster epididymis.
626-631
(1976).
A
surve" of sperm in the tubular Biol. Reprod. -15:
12.
Back, D.J. composition epididymis Experientia
and Shenton, J.C. A comparison of epididymal plasma from the of the rat, hamster and guinea (1975). -31:464-465
13.
Jones, R.C. spermatozoa elephant.
Luminal composition and maturation in the genital ducts of the African J. Repr0.d. Fertil. (1980). -60:87-93
1 G.
Setchell, B.P., Scott, T.W., Voglmayr, J.K. and Waites, G.M.H. Characteristics of testicular spermatozoa and the fluid which transports them into the epidldymis. Biol. Reprod. Suppl. 1: 40-60 (1969).
15.
Voglmayr, J.K., Waites, G.M.H. and Setchell, B.P. Studies on spermatozoa collected directly from the testis of the conscious rams. Nature 210:
861-863
16.
of the cauda pig.
of
(1964).
Voglmayr, J.K., Scott, T.W., qetchell, B.P. and Waites, G.M.H. Metabolism of testicular spermatozoa and characteristics of testicular fluid collscted from conscious rams. J. Reprod. Fertil.
-14:87-99
(1947).
NOVEMBER
1984 VOL. 22 NO. 5
THERIOCENOLOGY
17.
Wow,
18.
Kvist, IJ. Sperm nuclear chro‘natin decondensation ability. Acta. Physiol. Stand. Suppl. 436 : 1-24 (1980).
P.Y.D. tion in the epididymis.
NOVEMBER
Fluid reabsorpand Yeung, C.H. isolated duct of the rat cauda J. Reprod. Fertil. (1377). -43:77-81
1984 VOL. 22 NO. 5
551