Testicular Interstitial Fluid Volume Research Articles

Transient Rise of Serum Testosterone Level after Single Sildenafil Treatment of Adult Male Rats

Phosphodiesterase type 5 (PDE5) inhibitors have been established in therapy for a variety of physiological disorders including erectile dysfunction. Despite its popularity and wide usage in erectile dysfunction treatment, the short-term effect of PDE5 inhibition on Leydig cell functionality and testosterone dynamics is missing. This study was designed to assess the acute in vivo effects of sildenafil citrate (Viagra) treatment on testosterone production. Male adult rats were given sildenafil (1.25 mg/kg BW) per os, and testosterone production were analyzed 30, 60, 120, and 180 minutes after treatment. Additionally, in vitro effect of sildenafil extract on Leydig cell steroidogenesis was estimated. The formation of testicular interstitial fluid (TIF), and testosterone, cyclic guanosine monophosphate (cGMP), cyclic adenosine monophosphate (cAMP) content was followed. Occurrence and phosphorylation of mature steroidogenic acute regulatory protein (StAR) and interaction with protein kinase G 1 (PRKG1) were assessed by immunoprecipitation and Western blot. Serum testosterone was increased 60 and 120 minutes after sildenafil treatment. In 60 minutes, TIF volume was doubled and stayed increased till the end of the experimental period. cGMP and testosterone content in TIF were increased 30 minutes after treatment, and cAMP decreased in 60 minutes. Further, sildenafil-induced stimulation of testosterone production was abolished by ex vivo addition of PRKG1 inhibitor but not by protein kinase A inhibitor. Sildenafil treatment increased the level of phosphorylated and total StAR protein. Moreover, co-immunoprecipitation of StAR and PRKG1 was increased following sildenafil treatment suggesting the active role of this kinase in initiation of testosterone synthesis. Additionally, sildenafil extract applied in vitro on primary Leydig cell culture increased cGMP accumulation and testosterone production in time- and dose-dependent manner without effect on cAMP level. Acute sildenafil treatment enlarged TIF volume but also stimulated testosterone production which may be significant considering the positive testosterone effect in regulation of sexual activity.

Abstract: P172 THE INFLUENCE OF SILDENAFIL ON OXIDATIVE STRESS PARAMETERS DURING CORONARY AUTOREGULATION ON ISOLATED RAT HEART

Phosphodiesterase type 5 (PDE5) inhibitors have been established in therapy for a variety of physiological disorders including erectile dysfunction. Despite its popularity and wide usage in erectile dysfunction treatment, the short‐term effect of PDE5 inhibition on Leydig cell functionality and testosterone dynamics is missing.This study was designed to assess the acute in vivo effects of sildenafil citrate (Viagra) treatment on testosterone production.Male adult rats were given sildenafil (1.25 mg/kg BW) per os, and testosterone production were analyzed 30, 60, 120, and 180 minutes after treatment. Additionally, in vitro effect of sildenafil extract on Leydig cell steroidogenesis was estimated.The formation of testicular interstitial fluid (TIF), and testosterone, cyclic guanosine monophosphate (cGMP), cyclic adenosine monophosphate (cAMP) content was followed. Occurrence and phosphorylation of mature steroidogenic acute regulatory protein (StAR) and interaction with protein kinase G 1 (PRKG1) were assessed by immunoprecipitation and Western blot.Serum testosterone was increased 60 and 120 minutes after sildenafil treatment. In 60 minutes, TIF volume was doubled and stayed increased till the end of the experimental period. cGMP and testosterone content in TIF were increased 30 minutes after treatment, and cAMP decreased in 60 minutes. Further, sildenafil‐induced stimulation of testosterone production was abolished by ex vivo addition of PRKG1 inhibitor but not by protein kinase A inhibitor. Sildenafil treatment increased the level of phosphorylated and total StAR protein. Moreover, co‐immunoprecipitation of StAR and PRKG1 was increased following sildenafil treatment suggesting the active role of this kinase in initiation of testosterone synthesis. Additionally, sildenafil extract applied in vitro on primary Leydig cell culture increased cGMP accumulation and testosterone production in time‐ and dose‐dependent manner without effect on cAMP level.Acute sildenafil treatment enlarged TIF volume but also stimulated testosterone production which may be significant considering the positive testosterone effect in regulation of sexual activity. Janjic MM, Stojkov NJ, Bjelic MM, Mihajlovic AI, Andric SA, and Kostic TS. Transient rise of serum testosterone level after single sildenafil treatment of adult male rats. J Sex Med **;**:**–**.

Hormonal regulation and functional role of vascular endothelial growth factor a in the rat testis.

Vascular endothelial cell growth factor (VEGF-A) is synthesized in the testis but its role and regulation in this organ have not been examined. VEGF and its receptors (VEGF-R) were quantified using reverse transcription-polymerase chain reaction and Western blot. VEGF, VEGF-R1, and VEGF-R2 mRNAs and VEGF protein were increased after treatment with 50 IU hCG. Injection of 100 ng human recombinant VEGF 165 into the testis caused an increase in endothelial cell proliferation, but only a moderate increase in testicular interstitial fluid volume. In contrast with systemic hCG treatment, local VEGF injection did not increase the permeability to intravenously injected colloidal carbon particles. However, if VEGF was given locally in the testes of animals pretreated with hCG 4 or 8 h earlier, VEGF acted in synergy with hCG to increase vascular carbon leakage by forming interendothelial cell gaps. Testicular blood flow was unaffected by local VEGF 165 injection. Treatment with a specific VEGF-R2 tyrosine kinase inhibitor blocked the hCG-induced increase in endothelial cell proliferation but did not affect the hCG-induced accumulation of polymorphonuclear leukocytes in testicular blood vessels or the increase in the testicular interstitial space. The present study demonstrated that testicular VEGF secretion is increased by hormonal stimulation of Leydig cells and that VEGF, through effects mediated via VEGF-R2, regulates endothelial cell proliferation in the rat testis. VEGF does not appear to regulate testicular blood flow and it is not involved in inducing the hCG-induced inflammation-like response in the testicular microvasculature. The permeability-increasing effect of VEGF is low in the testis under basal conditions but is apparently up-regulated by hCG treatment.

Subchronic supplementation of lithium carbonate induces reproductive system toxicity in male rat

Lithium is frequently used as an effective drug for the treatment of several psychiatric disorders in human. This alkali element and its salt, at its higher doses, may lead to various side effects or has several toxic effects after prolonged therapeutic use. To test this hypothesis, the present study was designed to investigate the adverse effect of subchronic exposure of lithium carbonate on reproductive organs of male rat. Rats were exposed to lithium carbonate at doses of 500, 800, 1100 mg/kg of diet for 90 days. The weight of reproductive organs, histology of testis, epididymis, seminal vesicle, prostate, testicular interstitial fluid volume (IFV), testosterone level, sperm morphology and fertility index were analyzed. Treatment with higher doses of lithium carbonate (i.e. 800, 1100 mg/kg diet) significantly reduced testes, epididymis and accessory sex organs weights, whereas, lower dose (500 mg/kg diet) did not show any untoward effect. Similarly, the sperm number from cauda epididymis and daily sperm production was significantly decreased with higher doses of lithium carbonate. The serum testosterone levels and IFVs were also reduced significantly. Seminal vesicle and prostate secretions were completely blocked and spermatozoa were not seen in the lumen of epididymis and vas deference. Histological studies have revealed that lithium carbonate (1100 mg/kg) caused degeneration of spermatogenic cells and vacuolization of sertoli cells cytoplasm in the testis. The sperm transit rate and production of abnormal spermatozoa were significantly ( P<0.01) increased. When the lithium carbonate-treated males were mated with normal cyclic females, the fertility index declined to 50% even after 30 days of withdrawal of lithium carbonate treatment. These results clearly suggest that subchronic exposure of lithium carbonate promote reproductive system toxicity and reduces fertility of male rats.

Interactions Between Alcohol‐ and Opioid‐Induced Suppression of Rat Testicular Steroidogenesis In Vivo

To examine interactions between alcohol and endogenous opioids in their suppressive effects on rat testicular function, the opioid antagonist naltrexone or the opioid agonist morphine was administered to adult male rats alone or in combination with alcohol. Serum testosterone, testicular interstitial fluid (TIF) testosterone, and TIF volumes were measured to assess testicular function. Naltrexone induced dose-dependent increases in serum and TIF testosterone levels without changes in TIF volume. Alcohol (0.5 g/kg) inhibited naltrexone-induced stimulation of testosterone secretion and shifted the naltrexone dose-response curve to the right. Conversely, naltrexone (0.05 mg/kg) inhibited alcohol-induced suppression of testosterone secretion and shifted the alcohol dose-response curve to the right. Relatively high doses of naltrexone (5 to 30 mg/kg) were needed to stimulate testosterone secretion maximally in rats treated with a low dose of alcohol (0.5 g/kg) and to stimulate normal levels of testosterone secretion in rats treated with a high dose of alcohol (2 g/kg). In addition, combined treatment with 1 and 30 mg/kg of naltrexone and 0.5 to 2 g/kg of alcohol did not alter blood alcohol concentrations significantly, suggesting that the interactions between alcohol and naltrexone were unrelated to gross changes in alcohol metabolism or bioavailability factors. Simultaneous treatments with a low dose of alcohol (0.3 g/kg), near the threshold of efficacy, and low-moderate doses of morphine (0.3 to 3 mg/kg) were not additive in suppressing testosterone secretion, compared with either agent alone. These results support the hypothesis that opioid antagonists can reverse the suppressive effect of alcohol on testicular steroidogenesis, but the results also suggest that endogenous opioids do not exclusively mediate alcohol's effects on testosterone secretion.

Effect of short-term starvation on Leydig cell function in adult rats.

An experiment was carried out to analyze the effect of 3 days of starvation on the Leydig cell function in adult rats. Starvation markedly decreased plasma insulin and testosterone levels (p < .05). The weight of testes was maintained, whereas the testicular interstitial fluid volume decreased (p < .05). The level of testosterone decreased in this fluid (60%), whereas insulin levels showed no significant change. Purified Leydig cells showed normal LH/hCG binding in fasted rats but low insulin binding. The ability of these cells to produce testosterone in vitro was normal under both basal conditions and hCG stimulation in the absence and presence of insulin in the incubation medium. These data suggest that there is no gross impediment in the Leydig cell capacity to produce testosterone that might explain the starvation-associated decrease in plasma testosterone.

Effect of a Dopamine agoniste on the development of Leydig cellhyperplasia in Sprague-Dawley rats

Dopamine agonists are known to increase the incidence of Leydig cell hyperplasia/adenomas when administered to rats over periods of 1–2 years. We have examined the early changes in factors affecting Leydig cell growth/hyperplasia after chronic oral administration of one of these dopamine agonists, Mesulergine (CU32-085) N-(1-6-dimethylergolin-8α-yl)-N,N-dimethylsulphamide hydrochloride), to Sprague-Dawley (SD) rats. Eight-week-old rats were given the dopamine agonist (2 mg/kg body weight/day) in food for 5 or 57 weeks. The dopamine agonist treatment had no significant effect on food intake, body weight, and testis and seminal vesicle size, but significantly decreased testicular interstitial fluid volume at 5 weeks (by 51%). Leydig cells isolated from rats treated with the dopamine agonist for 5 weeks exhibited an increase in the rate of protein synthesis compared with the controls (by 28%). This treatment, however, had no significant effect on the number of Leydig cells or macrophages as assessed by histological examination of testicular sections. Treatment with the dopamine agonist for 57 weeks caused a 36 and 28% increase in the number of Leydig cells and macrophages, respectively. Nodules of Leydig cells, indicating the first signs of tumor development, were present in testes from the 57- but not the 5-week-treated animals or the controls of both groups, although an increase in the number of Leydig cells occurred with aging. Thick-walled arterioles were found in the intertubular spaces of the testis sections from rats treated for 57 weeks. These findings suggest that chronic treatment of male SD rats with the dopamine agonist causes hypertrophy of Leydig cells within 5 weeks (as assessed by [3H]methionine incorporation), followed by hyperplasia within 2 years, prior to the development of Leydig cell adenomas, which occur within 1–2 years after the initiation of treatment.

Effect of a dopamine agonist on the development of Leydig cell hyperplasia in Sprague-Dawley rats.

Dopamine agonists are known to increase the incidence of Leydig cell hyperplasia/adenomas when administered to rats over periods of 1-2 years. We have examined the early changes in factors affecting Leydig cell growth/hyperplasia after chronic oral administration of one of these dopamine agonists, Mesulergine (CU32-085) [N-(1-6-dimethylergolin-8 alpha-yl)-N,N-dimethylsulphamide hydrochloride), to Sprague-Dawley (SD) rats. Eight-week-old rats were given the dopamine agonist (2 mg/kg body weight/day) in food for 5 or 57 weeks. The dopamine agonist treatment had no significant effect on food intake, body weight, and testis and seminal vesicle size, but significantly decreased testicular interstitial fluid volume at 5 weeks (by 51%). Leydig cells isolated from rats treated with the dopamine agonist for 5 weeks exhibited an increase in the rate of protein synthesis compared with the controls (by 28%). This treatment, however, had no significant effect on the number of Leydig cells or macrophages as assessed by histological examination of testicular sections. Treatment with the dopamine agonist for 57 weeks caused a 36 and 28% increase in the number of Leydig cells and macrophages, respectively. Nodules of Leydig cells, indicating the first signs of tumor development, were present in testes from the 57- but not the 5-week-treated animals or the controls of both groups, although an increase in the number of Leydig cells occurred with aging. Thick-walled arterioles were found in the intertubular spaces of the testis sections from rats treated for 57 weeks. These findings suggest that chronic treatment of male SD rats with the dopamine agonist causes hypertrophy of Leydig cells within 5 weeks (as assessed by [3H]methionine incorporation), followed by hyperplasia within 2 years, prior to the development of Leydig cell adenomas, which occur within 1-2 years after the initiation of treatment.

Evaluation of the impression technique by measuring interstitial oedema in rat testis

The impression method for assessment of subcutaneous oedema was evaluated in a rat testis model where the testicular interstitial fluid volume was changed both artificially by infusion of rat plasma and pharmacologically by administration of human chorionic gonadotrophin. Both the integral value and the impression force value, as measured with the impression method, changed with infused volume, and changes as small as 16 microliters (approximately 7% of the total interstitial fluid volume in a testis) could be detected. Rats were treated with a single injection of 100 i.u. human chorionic gonadotrophin in order to induce changes in the volume of interstitial fluid in the testis. Both the integral value and the impression force value seemed to reveal information on testicular interstitial fluid volume in rats treated with human chorionic gonadotrophin that was similar to data revealed by measuring the actual fluid content in the testis. Interstitial fluid volume measured morphometrically in the contralateral testis in human chorionic gonadotrophin-treated rats was significantly correlated to the impression force value (r = 0.75) and the integral value (r = 0.52). This rat testis model proved to be an interesting experimental set-up for evaluation of the impression technique.

Effects of Alcohol on β‐Endorphin and Reproductive Hormones in the Male Rat

In an attempt to examine the relationship between alcohol-induced alterations in immunoreactive beta-endorphin (i-beta E) levels in the hypothalamic-pituitary-gonadal axis and the synthesis and release of reproductive hormones, male rats were treated with either an acute intraperitoneal injection of alcohol or were chronically exposed to an alcohol-containing liquid diet. Hypothalamic, pituitary, serum, and testicular levels of immunoreactive beta-endorphin (i-beta E) and serum levels of luteinizing hormone (LH) and testosterone were measured at various times after initiation of these treatments. Testicular interstitial fluid (TIF) volumes and levels of TIF i-beta E and testosterone were also measured 4 hr after acute treatment as an index of testicular release of these substances. Acute alcohol decreased pituitary levels of i-beta E and increased serum levels of the peptide for up to 1 hr after its injection, but did not alter hypothalamic or testicular levels. Acute alcohol markedly increased TIF i-beta E and decreased TIF testosterone and TIF volume. Sharp decreases in serum LH and testosterone were observed in association with these acute changes in i-beta E levels in the pituitary, blood, and testes. During chronic alcohol exposure serum testosterone levels were substantially depressed, but tolerance appeared to develop quickly to the chronic effects of alcohol on serum LH. Similarly, tolerance to alcohol's effects on i-beta E levels in the pituitary and serum also appeared to develop during chronic alcohol administration. However, hypothalamic and testicular i-beta E levels were markedly suppressed by chronic alcohol administration in contrast to the lack of effect observed after acute alcohol administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Evidence for the control of testicular interstitial fluid volume in the rat by specific germ cell types

Following on from our recent evidence that Sertoli cells may regulate testicular interstitial fluid (IF) volume, this study has assessed whether depletion of specific germ cell types in vivo is associated with changes in recovered IF volume. Germ cell depletion was induced by either a single oral administration of 650 mg methoxyacetic acid (MAA)/kg or exposure of the testes to local heating (43 degrees C for 30 min). Treatment with MAA induced depletion or loss of most pachytene and later spermatocytes at 1-3 days and, because of maturation depletion, this resulted in the specific depletion of later germ cell types at 7-35 days. Testicular IF volume was unchanged at 1-7 days after MAA treatment but was increased significantly (P less than 0.01) at 14 days and was nearly doubled (P less than 0.001) at 21 days, before returning to control levels at 28-42 days. Serum LH (and FSH) levels were generally higher in MAA-treated rats, especially at 21 and 28 days, but there was no obvious correlation between LH levels and IF volume changes. Similarly, there was no relationship between IF volume changes and testicular weight or IF levels of testosterone. The increase in IF volume at 14-21 days after MAA treatment coincided with specific depletion of the later elongate spermatids (steps 14-19) and, when these cells reappeared in the testis, IF volume normalized. This possible causal association was studied further in rats exposed to local testicular heating which, within 3 days, caused major depletion of pachytene spermatocytes and early (step 1-8) spermatids.(ABSTRACT TRUNCATED AT 250 WORDS)

High and low doses of luteinizing hormone induce different changes in testicular microcirculation.

To study the effects of LH on total testicular blood flow and microcirculation, rats were treated with 2.5 or 25 micrograms LH sc and measurements were made on control rats and on LH-treated rats 1, 2, 3, 4, 6 and 12 h, after treatment. After treatment with 25 micrograms LH, total testicular blood flow, as measured with the radioactive microsphere method, was decreased at 6 h and increased at 12 h. Testicular microcirculatory blood flow was recorded with laser Doppler flowmetry and regular oscillations in blood flow, vasomotion, was observed in control rats. Vasomotion was not present 4 and 6 h after treatment with 25 micrograms LH, but returned at 12 h. Prior to and concomitantly with these changes in vasomotion, polymorphonuclear leukocytes accumulated in testicular microvessels and migrated into the interstitial tissue. These changes were followed, 6 h after treatment, by an increased vascular permeability, measured as increased testicular interstitial fluid volume. The lower dose of LH (2.5 micrograms), doubled plasma testosterone concentration and initially decreased interstitial fluid volume, and later induced a slight increase in blood vessel leukocytes. At the times studied, no changes could be observed in the other vascular parameters. In conclusion, it is suggested that LH, probably via some Leydig cell product, promotes regulatory effects on testicular microcirculation, but different magnitudes of LH stimulation induce different responses.

Effect of an LHRH-agonist on testicular microcirculation in hypophysectomized rats.

The effect of an LHRH-agonist on testicular vasomotion was studied in adult hypophysectomized rats. At 0.5 and 1 h after injection of 1 microgram LHRH-agonist, no effect on vasomotion was observed, but after 2 h the pulsatile blood flow pattern had changed to continuous flow in 50% of the rats. At 4 h after treatment with LHRH-agonist, vasomotion was not detected in the testis of any rat. In addition, at the latter time an increased number of polymorphonuclear leucocytes (PMNs) were observed in testicular microvessels, and some PMNs had migrated into the interstitial space. These findings are discussed in relation to the well-known stimulatory effect of LHRH-agonists on testicular interstitial fluid volume.

Testicular Blood Flow and Microcirculation in Rats After Treatment with Ethane Dimethyl Sulfonate1

The effects of ethane dimethyl sulfonate (EDS) on total testicular blood flow, microcirculation, and the testicular interstitial fluid volume (IFV) in rats were studied. In agreement with previous studies, treatment of control rats with human chorionic gonadotropin (hCG) induced an increase in IFV and total testicular blood flow as measured with radioactive microspheres. These effects of hCG were completely abolished in rats pretreated with EDS; in EDS-treated rats not receiving any hCG, there were decreases in IFV when compared with untreated control rats. Furthermore, the pulsatile pattern of testicular microcirculation registered with laser-Doppler flowmetry was abolished after EDS treatment, and this effect was not influenced by hCG treatment. The hCG-induced increase in IFV is associated with an increased accumulation of polymorphonuclear leukocytes locally in the testis, but this accumulation of leukocytes was not observed in rats pretreated with EDS. It was concluded from the present study that hCG-induced changes in total testicular blood flow and testicular microcirculation require functionally intact Leydig cells.

Effects of oestradiol-17 beta on testicular microcirculation in rats.

The effect of oestradiol-17 beta on testicular microcirculation in intact and hypophysectomized rats was studied before and after treatment with human chorionic gonadotrophin (hCG). Treatment of intact rats with oestradiol-17 beta for 5 days did not influence vasomotion but decreased testicular interstitial fluid volume (IFV). Treatment of intact rats with 50 IU hCG 8 h before the experiment began induced an increase in testicular IFV, abolished vasomotion and increased the accumulation of polymorphonuclear leucocytes in the testicular venules and interstitium. These changes were unaffected by pretreatment with oestradiol-17 beta, despite the decreased testosterone production. However, pretreatment with oestradiol-17 beta potentiated the hCG-induced migration of polymorphonuclear leucocytes to the interstitium. The interstitial fluid volume and number of polymorphonuclear leucocytes in blood vessels were decreased in hypophysectomized rats, and vasomotion was abolished. Daily treatment with 5 IU hCG increased the IFV and the number of polymorphonuclear leucocytes in blood vessels, and preserved vasomotion. Treatment of hypophysectomized rats with oestradiol-17 beta decreased testosterone production but did not influence basal IFV, vasomotion or the changes in IFV and vasomotion induced by 5 IU hCG. The present study shows that the regulation of testicular vascular permeability and vasomotion may not be directly related to testicular steroidogenesis, and that oestrogens are probably not involved as a mediator of the hCG-induced changes in testicular microcirculation.

Leucocytes mediate the hCG-induced increase in testicular venular permeability

Human chorionic gonadotrophin (hCG) treatment of adult male rats induces microvascular changes in the testis consisting of abolished vasomotion, an accumulation of leucocytes in postcapillary venules and increased vascular permeability. To study the role of leucocytes, rats were made leucopenic with a specific antineutrophil serum (ANS). Testicular interstitial fluid volume was decreased in leucopenic rats. Leucopenic rats also failed to show an hCG-induced increase in venular permeability as in saline-reated rats. The normally pulsatile blood flow pattern (vasomotion) persisted in leucopenic rats but was abolished after hCG treatment both in saline-treated and leucopenic rats. Plasma testosterone concentration after hCG treatment was not affected by elimination of circulating polymorphonuclear (PMN) leucocytes. It is concluded that PMN leucocytes mediate in part the hCG-induced increase in testicular venular permability but not the hCG-induced inhibition of vasomotion.

Treatment of rats with hCG induces inflammation-like changes in the testicular microcirculation.

Adult rats were injected subcutaneously with 50 i.u. hCG and vascular permeability was compared to that in saline-treated control rats by two independent methods. At 4 h after hCG treatment the rats were injected intra-arterially (i.a.) with FITC-labelled macromolecular dextran (Mr 150,000) and the testicular microcirculation was studied in vivo by using a fluorescence microscope. Other rats were injected i.a. with a suspension of colloidal carbon and the location of leaking blood vessels was recorded in sections from the testes by light and electron microscopy. In hCG-treated animals leucocytes were found adhering to the endothelium in post-capillary venules and in these venular segments dextran was leaking into the interstitium. Carbon particles were deposited in the walls of post-capillary venules and leucocytes migrated through open interendothelial cell gaps in hCG-treated animals. In control animals leucocyte adhesion and migration were not observed, the injected dextran remained in the circulation and the blood vessels were not labelled by carbon. It is suggested that the hCG-induced increase in testicular interstitial fluid volume, like the tissue oedema in inflammation, is caused by a leucocyte-mediated increase in venular permeability.

Relationship between human chorionic gonadotrophin-induced changes in testicular microcirculation and the formation of testicular interstitial fluid.

The relationship between testicular vascular permeability and testicular microcirculation as measured by laser Doppler flowmetry was studied in adult rats. In untreated control animals there was an oscillatory testicular blood-flow pattern with a frequency of 10.6 +/- 0.8 pulses/min and the amount of testicular interstitial fluid (IF) collected was 61.5 +/- 2.2 microliter/g testis. Treatment of the rats with 25-200 i.u. human chorionic gonadotrophin (hCG) s.c. 8 h before the experiment resulted in a change in the testicular flow pattern from pulsatile to continuous and an increase in IF volume. Treatment with hCG (50 i.u., s.c.) changed the testicular blood-flow pattern from oscillatory to continuous 4, 8 and 16 h after treatment. The flow pattern returned to being pulsatile 32 h after treatment with hCG. The IF information was increased at those times when the blood-flow pattern was continuous. No effects on blood flow or IF formation were observed with 12.5 i.u. hCG s.c. The present study shows a dose- and time-dependent covariation between the increase in testicular IF volume and the disappearance of the pulsatile flow in testicular microcirculation. It appears that a continuous flow pattern favours the transport of fluid from blood vessels to the interstitium.

Proopiomelanocortin-derived peptides in testicular interstitial fluid: characterization and changes in secretion after human chorionic gonadotropin or luteinizing hormone-releasing hormone analog treatment.

Recent studies indicate that proopiomelanocortin (POMC)-derived peptides are present in the testis and may be involved in the regulation of gonadal function, although their precise role still remains obscure. In the present study, we investigated in the rat whether testicular interstitial fluid (TIF), a functionally important compartment of the testis, contains POMC-derived peptides. Adult rats were used for all the experiments, and TIF was collected from each individual testis and analyzed for the presence of ACTH and beta-endorphin-like immunoreactivity (beta end-LI). Initial studies indicated that both ACTH and beta end-LI can be readily detected in TIF from intact rats, and that the concentrations of these peptides are severalfold higher than those in the peripheral plasma of the same animals. Similar studies in rats 3-5 days after hypophysectomy indicate that although ACTH and beta end-LI levels in plasma were undetectable, high levels of both peptides were measured in TIF. Moreover, serial dilution curves of TIF showed parallelism with the respective standard curves in the beta end and ACTH assays. Additional studies indicated that levels of POMC peptides in TIF were modified by treatments that affect the endocrine function of the testis. In that respect, hCG given sc to hypophysectomized rats increased testosterone levels in TIF and plasma, and similarly increased beta end-LI concentrations in TIF. A LHRH analog (LHRH-A; [D-Ala6,des-Gly10]LHRH-ethylamide) given sc to hypophysectomized rats resulted in increased testosterone production, but decreased beta end-LI in TIF, suggesting that the effects of hCG and LHRH-A on testicular beta end-LI secretion are not directly coupled with testosterone production. Prolonged treatment of intact rats with the LHRH-A for 2-10 days resulted in progressive declines in testicular weight and testosterone levels in TIF and in plasma, and a significant suppression of beta end-LI levels in TIF as early as 2 days after analog treatment. On the other hand, acute intratesticular injection of LHRH-A to intact rats (5 ng into the right testis) produced a short-lived increase in beta end-LI in TIF 2 h after injection, coincident with a temporary decrease in TIF volume. These results indicate that the POMC-derived peptides ACTH and beta end are present in TIF and are secreted locally into this compartment, supporting previous reports demonstrating the presence and local synthesis of POMC peptides in testicular tissue.(ABSTRACT TRUNCATED AT 400 WORDS)

Factors determining whether the direct effects of an LHRH agonist on leydig cell function in vivo are stimulatory or inhibitory

The influence of (1) repeated exposure to LHRH agonist and (2) concomitant exposure to various levels of LH or hCG, on the direct testicular effects of LHRH agonist have been investigated in hypophysectomized and intact rats. In the latter, LHRH agonist was injected intratesticularly into the right testis whilst the left testis was injected with vehicle, and the level of testosterone in interstitial fluid (IF) from left and right testes was compared 2-4 h later. In hypophysectomized rats, a single injection of 50-1000 ng LHRH agonist raised (P less than 0.001) serum levels of testosterone 2 h later to within the normal range (1-8 ng/ml) for intact rats of comparable age. Subsequent daily injections of LHRH agonist elicited progressively smaller responses and by day 5 no response was evident, this decline being unrelated to the increase in time after hypophysectomy. Comparable changes were observed in hypophysectomized rats pretreated with an LH antiserum. Daily injection of hypophysectomized rats with 10 IU hCG for 6 days raised serum levels of testosterone to supraphysiological levels on each day, whilst concomitant injection of LHRH agonist (1 microgram) decreased (P less than 0.001) this response at all times, an effect that became progressively more pronounced. In contrast, daily treatment with 1 microgram ovine LH raised serum levels of testosterone to within the physiological range, and concomitant treatment with LHRH agonist (50 ng) had either no effect (days 1-2) or significantly increased (days 4-6) the testosterone response to LH. In intact rats, a single unilateral intratesticular injection of 1 ng LHRH agonist increased the IF levels of testosterone unilaterally 3 h later, but subsequent injections on days 2 and 3 elicited progressively smaller responses. In rats given a single intratesticular injection of LHRH agonist combined with a peripheral injection of different doses of LH or hCG, the LHRH agonist induced a unilateral increase in IF levels of LH/hCG, whilst in rats treated with high doses of LH (12.5--25 micrograms) or hCG (50 IU). LHRH agonist either had no effect or significantly reduced the IF levels of testosterone unilaterally. However, LHRH agonist also had significant effects on testicular IF volume and, as this may reflect altered transport of LH and hCG to the Leydig cells, the inhibitory effects of LHRH agonist may be related to this change rather than to an effect of steroidogenesis itself.(ABSTRACT TRUNCATED AT 400 WORDS)