Day Way Goh

Prenatal androgen blockade with flutamide inhibits masculinization of the genitofemoral nerve and testicular descent

Prenatal androgen blockade with the antiandrogen flutamide inhibits the inguinoscrotal phase of testicular descent. The evidence suggests that androgens may act indirectly via the sexually dimorphic genitofemoral nerve (GFN) to control this phase. Rats were exposed to flutamide on gestational days 16 through 19. Seven-day-old rats were subjected to retrograde fluorescent labelling of the GFN combined with immunohistochemistry for calcitonin gene-related peptide (CGRP), a neurotransmitter found in the GFN. Fluorescent-labelled and CGRP-immunoreactive neurons in the GFN spinal nucleus were quantified. Sexual dimorphism of the GFN nucleus was absent in the flutamide-treated rats but obviously present in control rats. Furthermore, control male nuclei had 24% more CGRP-immunoreactive neurons and 12% more fluorescent-labelled neurons than did flutamide-treated male nuclei. This study shows that prenatal androgen blockade with flutamide inhibits masculinization of the GFN, with significant reduction of its CGRP content. This supports the proposal that androgens act via the GFN, with CGRP as the second messenger, to control inguinoscrotal testicular descent.

The relationship among calcitonin gene-related peptide, androgens and gubernacular development in 3 animal models of cryptorchidism

The relationship among calcitonin gene-related peptide (CGRP), a neurotransmitter in the genitofemoral nerve, androgens and gubernacular development was studied using rats treated prenatally with the antiandrogen flutamide and the mutant cryptorchid TS rat. We compared these 2 groups with the testicular feminization mouse with androgen insensitivity. Gubernacula from male TS rats and flutamide-treated rats were maintained in organ culture and examined for contractile response to CGRP. Controls were gubernacula from normal rats and vehicle-treated rats, respectively. TS rat gubernacula have an inhibited contractile response to CGRP, whereas flutamide-treated rat gubernacula have an exaggerated response. A similar exaggerated response to CGRP has previously been demonstrated in testicular feminization mouse gubernacula. These results revealed abnormalities in gubernacular contractile response to CGRP in these cryptorchid animal models, implying that CGRP and gubernacular contractility may have key roles in mediating normal inguinoscrotal testicular descent.

The effect of flutamide on testicular descent in rats examined by scanning electron microscopy

The effect of prenatal flutamide exposure on testicular descent was investigated by using scanning electron microscopy (SEM) in prenatal and postnatal rats. In 20-day-old fetal rats, SEM showed no significant difference in the degree of gubernacular development or testicular descent relative to the kidney between flutamide-treated (74.5 +/- 2.2 U) and control rats (73.3 +/- 1.5 U); however, there was significant inhibition in oestrogen-treated rats (44.3 +/- 2.2 U) (P < .001). (The distance between the kidney and the bladder neck was standardized to 100 U.) In 5-day-old rats, SEM showed inhibited downward growth of the processus vaginalis in flutamide-treated rats. The length of processus vaginalis below the inguinal ligament was 32.8 +/- 2.4 U in flutamide-treated rats and 51.7 +/- 1.8 U in controls (P < .001). In 30- to 35-day-old mature rats, the frequency of cryptorchidism was 41.3% for flutamide-treated rats and 0% for controls (P < .001). Some cryptorchid testes were located in the lower abdominal cavity (10.9%); others were in the suprainguinal position (26.1%) or on the line of descent in the inguinal region (4.3%). In the flutamide-treated group, no testes were located in the posterior abdominal cavity, near the kidney. These results suggest that transabdominal descent of the testis is independent of androgen action, but that androgens control inguinoscrotal descent of the testis by regulating gubernacular migration and the growth of the processus vaginalis.

Is the retractile testis a normal, physiological variant or an anomaly that requires active treatment?

Conventional teaching states that the retractile testis is a normal, physiological variant that descends spontaneously by puberty and requires no active treatment. Critical review of the literature, however, suggests that this complacent view may be inappropriate. Substantial overlap exists between the three seemingly separate entities of the late descending, the ascending and the retractile testes. This overlapping group probably accounts for the recently observed increased incidence of orchidopexies. Retractile testes that spend most of the time in an extrascrotal position are subject to the same adverse effects of higher temperatures as “true” undescended testes, regardless of whether they can be manipulated into the scrotum; what matters is where they actually reside most of the time. The evidence suggests that such retractile testes suffer similar pathological changes to “true” undescended testes if left to await spontaneous descent. Evidence is presented to support a radical, novel proposal that the retractile testis is a variant of the spectrum of pathological maldescended testes and requires active treatment. A new strategy is proposed for the management of this common pathology.

Calcitonin Gene-Related Peptide Receptors in the Gubernaculum of Normal Rat and 2 Models of Cryptorchidism

We investigated calcitonin gene-related peptide binding in the gubernaculum from a normal rat, a TS rat (congenitally cryptorchid from unknown causes) and a flutamide rat (cryptorchid secondary to prenatal antiandrogen treatment). Gubernacular sections from 2-day-old male pups were incubated with 125iodine-calcitonin gene-related peptide with various concentrations of unlabeled calcitonin gene-related peptide. After exposure to x-ray film, developed film was quantified by computerized densitometry. The binding analysis showed that concentrations of calcitonin gene-related peptide receptors were 22.9 +/- 2.12, 13.5 +/- 1.70 and 30.3 +/- 2.63 (mean plus or minus standard error of mean, fmol./mg. polymer) for each rat, respectively, and there were significant differences between the normal and TS rats (p < 0.01), and the normal and flutamide rats (p < 0.05). However, there was no significant difference in the dissociation constant among the 3 models. This result shows that deficiency and excess of calcitonin gene-related peptide binding in the gubernaculum are associated with cryptorchidism. Calcitonin gene-related peptide released from the genitofemoral nerve may regulate its own receptor concentration to control gubernacular motility in vivo.

Calcitonin gene-related peptide stimulates motility of the gubernaculum via cyclic adenosine monophosphate

Calcitonin gene-related peptide (CGRP), N-truncated CGRP fragments CGRP 8-37 and [Tyr0]-CGRP 28-37, and dibutyryl cyclic adenosine monophosphate (DBcAMP) were studied for their effects on the neonatal male mouse gubernaculum in organ culture. Rhythmic contractions were shown in 18% of control gubernacula, which were enhanced with CGRP, inhibited by CGRP 8-37 and not affected by [Tyr0]-CGRP 28-37. A total of 60 gubernacula was exposed to increasing concentrations of DBcAMP and the percentage of gubernacula showing rhythmic contractions increased from 18 to 60%. These studies demonstrate that the neonatal mouse gubernaculum exhibits endogenous contractility that can be enhanced with CGRP or DBcAMP. These results suggest that cyclic adenosine monophosphate may act as the intracellular second messenger for receptor bound CGRP in the gubernaculum. This finding is consistent with the hypothesis that CGRP from the genitofemoral nerve provides directional, chemotactic guidance for inguinoscrotal gubernacular migration during testicular descent.

The Role of the Genitofemoral Nerve and Calcitonin Gene-Related Peptide in Congenitally Cryptorchid Mutant TS Rats

We investigated whether previous gubernacular denervation in TS rats changes the gubernacular contractile response to or binding capacity of calcitonin gene-related peptide receptors. In TS rats there is excessive calcitonin gene-related peptide in the genitofemoral nerve and 85% of the male rats have cryptorchidism. TS rats and controls underwent genitofemoral nerve transection or sham operation at day 0. On day 3 gubernacula were cultured with or without calcitonin gene-related peptide to observe contractions. Gubernacular frozen sections were incubated with 125iodine calcitonin gene-related peptide with or without unlabeled calcitonin gene-related peptide. Exposed x-ray films were quantified by computerized densitometry. In the sham operated controls contractility increased from 15 to 55% by adding calcitonin gene-related peptide compared with 5% and 10%, respectively, in the TS rats. After genitofemoral nerve denervation exogenous calcitonin gene-related peptide increased contractility to 90% in controls and 75% in TS rats. Previous genitofemoral nerve transection increased specific 125iodine calcitonin gene-related peptide binding 21% in controls and about 100% in TS rats. Genitofemoral nerve transection in TS rats restored gubernacular contractile response to calcitonin gene-related peptide in vitro and doubled the binding capacity of calcitonin gene-related peptide receptors.

Physiological effects in vitro of calcitonin gene-related peptide on gubernacular contractility with or without denervation

The gubernaculum in neonatal rats has been shown previously by direct observation to contract rhythmically in response to exogenous calcitonin gene-related peptide (CGRP), but the physiological properties of these contractions were unknown. In the first study the authors investigated gubernacular contractility in vitro using a strain gauge to see if there were characteristics of skeletal or smooth muscle. Both the frequency and the amplitude of contractions were significantly enhanced by CGRP, and isotonic tension of the gubernaculum and the duration of contractions were also increased after CGRP. The effect of CGRP on gubernacular contractions appeared several minutes after adding CGRP, and it was independent of the acetylcholine action, which induced only a single twitch response of the gubernaculum. In the second study the authors investigated the effect on gubernacular contractility of prior transection of the genitofemoral nerve (GFN), which contains CGRP. Vigorous contractions occurred in 85% of denervated gubernacular compared with 46% of controls (P < .01). These data provide the first quantitative evidence of rhythmic gubernacular contractions, and suggest that CGRP enhances gubernacular contractility by a direct effect independent of acetylcholine. Further, the contractile properties resemble those of differentiated cardiac muscle or primitive embryonic skeletal muscle. GFN transection enhances the gubernacular contractile response to exogenous CGRP, which is consistent with the GFN being the normal source of CGRP for the gubernaculum in vivo.

Portal hypertension in children : the changing spectrum

This report is an analysis of changing trends in the etiology and management of portal hypertension (PHT) in childhood. The study compared the 53 admissions to the Royal Childrens Hospital from 1971 to 1991 (33 intrahepatic, 20 extrahepatic) with the 77 admissions (22 intrahepatic, 54 extrahepatic, 1 with a congenital anomaly of the mesenteric vein) from the previous 23-year period (1948 to 1971). In addition to the differences in etiology, there has been (1) an increasing role for sclerotherapy as a therapeutic modality (and with this, lessening of the role of surgery), (2) a different approach to investigation (particularly imaging techniques), and (3) the availability of organ transplantation.

Exogenous calcitonin gene-related peptide causes gubernacular development in neonatal (Tfm) mice with complete androgen resistance☆☆☆

It has been proposed that testicular descent is controlled indirectly by androgens acting on the central nervous system to mediate migration of the gubernaculum to the scrotum. Accumulating evidence suggests that the genitofemoral nerve may release a newly described neurotransmitter, calcitonin gene-related peptide (CGRP) to stimulate gubernacular motility during migration. This study aimed to determine whether exogenous CGRP could stimulate gubernacular migration in mice with complete androgen resistance (testicular feminization mouse [Tfm]). CGRP was injected into the right groin of neonatal Tfm mice at 2-day intervals until 2 weeks of age, when the length of the processus vaginalis was measured under a dissecting microscope. The processus vaginalis length in normal male littermates was 5.9 +/- 1.8 mm (mean +/- SD) while in the female it was 1.2 +/- 0.9 mm. Exogenous CGRP had no effect on either of these. In Tfm males CGRP caused a significant increase in the length of the processus vaginalis on the injected side (2.3 +/- 0.8 mm) compared with the uninjected side (1.4 +/- 1.0 mm). These results are consistent with the hypothesis that CGRP can replace, at least partially, the effect of androgens on gubernacular migration.