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Summary. The secretory activity of the subcommissural organ (SCO) is affected by adrenalectomy, adrenalectomy + castration, and by an increase in ambient.
Cell Tissue Res. 200, 323-327 (1979)

Cell and Tissue Research 9 by Springer-Verlag 1979

Short Communication

The Subcommissural Organ of Lacerta s. sicula Raf.: Functional Studies* V. D'Uva, G. Ciarcia and A. Ciarletta Institute of Histology and Embryology,Laboratory of ComparativeAnatomy, Naples, Italy

Summary.The secretory activity of the subcommissural organ (SCO) is affected by adrenalectomy, adrenalectomy + castration, and by an increase in ambient temperature in adrenalectomized and adrenalectomized + castrated animals. Adrenalectomy inhibits the activity of the SCO. After adrenalectomy + castration the decrease in the secretory activity of the SCO is more rapid. In contrast, an increase in the ambient temperature in adrenalectomized animals induces a recrudescence of the activity of the SCO. The increase in temperature in castrated + adrenalectomized lizards does not affect the inhibition produced by this type of surgical treatment. The histological changes are discussed on the basis of results obtained in the present study and in previous experiments. Key words: Subcommissural organ - Adrenalectomy - Castration - Lacerta s. sicula Raf. (Lacertilia).

The subcommissural organ (SCO) of Lacerta s. sicula Raf., a secretory ependymal specialization located in the region of the posterior commissure, has been investigated for several years in this laboratory (D'Uva and Ciarcia, 1976; D'Uva et al., 1976, 1977, 1978; Varano et al., 1978). These studies indicate that the secretory activity of the SCO of Laeerta s. sieula shows seasonal variations. On the basis of light and electron microscopic changes, it has been possible to recognize four distinct stages of the secretory cycle of this organ: The first stage, typical of December, is characterized by a reduced secretory activity. The cells of the SCO contain only a few secretory granules located near the apical pole (type A and B granules). The basal portion of the cell is almost devoid o f granular material. The second stage, found from January to March and from midSeptember to November, is characterized by the presence of large masses of Send offprint requests to: Dr. Vittorio D'Uva, Universit/tdegli Studi di Napoli, Istituto di Istologia ed

Embriologia, Via Mezzocannone, 8, 80134 Napoli, Italy * This work was presentedat the "First Colloquiumof the EuropeanPineal StudyGroup", November 20-24, 1978, in Amsterdam

0302-766X/79/0200/0323/$01.00

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secretory material in the basal portion of the cells. Electron microscopically, these inclusions are located in cisternae of the rough endoplasmic reticulum; they consist of a material of low electron density (type C secretory material). The third stage, found from March to the end of May and from July to mid-September, is characterized by an increase in the number of the small granules and by the presence of large masses of secretory material, also found in the supranuclear region of the cells (type C secretory material). During the fourth stage, which is reached in June, the cells are filled with various types of secretory material (type A and B granules, type C secretory material). Further investigations, carried out in an attempt to elucidate the existence of possible relationships between the steroidogenic activity of the testes and the secretion of the SCO (D'Uva et al., 1977), point out that the SCO is influenced by all the experimental procedures that stimulate the Leydig cells and thus the secondary sexual characteristics (SSC). In contrast, all experiments that reduce or suppress the production of testicular steroids (castration, cyproterone treatment) are very effective in reducing the activity of the SCO. Furthermore, in the lizard Lacerta s. sicula a study of the possible relationship between the activity of the adrenal gland and the subcommissural organ demonstrates that the corticosteroids alone are capable of affecting the activity of the SCO (Varano et al., 1978). Data obtained from other experiments suggest that ambient temperature also plays an important role in the morphological changes occurring in the SCO after exogenous hormone administration. Only above an ambient temperature of 12~C, testosterone administration has a stimulating effect upon the secretory activity of the SCO. On the other hand, both in summer and winter animals, the activity of the SCO can be inhibited after suppression of the androgen action either by castration or by administration of cyproterone. Thus, both humoral and thermic factors are involved in the control of the SCO. In order to analyze the role of steroid hormones and temperature in the regulation of the secretory activity of the SCO in Lacerta s. sicula Raf., it appeared suitable to study (i) the effects of adrenalectomy and castration, and (ii) the influence of high temperatures in adrenalectomized, castrated and cyproteronetreated animals.

Materials and Methods A total of 180 adult males o f L a c e r t a s. sicula Raf., captured in March and May, were used for the present study. These animals were (i) adrenalectomized, or (ii) castrated + adrenalectomized immediately after their capture. A group of adrenalectomized and a second group of castrated + adrenalectomized lizards were transferred one week after the operation into a special room at 28 ~C, where they were kept under natural photoperiod with an air humidity of 65 %. Other groups were kept under natural conditions of temperature, photoperiod and humidity. A n u m b e r of castrated + adrenalectomized animals were treated with 100 gg testosterone, administered intraperitoneally in 0.5 cc almond oil (one injection every three days). The treatment with testosterone was started one week after castration + adrenalectomy. A n o t h e r group of lizards was treated in May with 0.5 m g cyproterone. The experimental and sampling procedures are summarized in Table 1. The animals were fed on larvae of Tenebrio molitor and fresh fruit ad libitum. The brains were fixed in Stieve's fluid and embedded in paraffin-celloidin according to Peterfi; 6 lam thick sections were stained with chromalum-hematoxylin and phloxin according to Gomori-Bargmann.

Subcommissural Organ of Lacerta s. sicula Raf.

325

Results and Discussion

Group A: Controls. The SCO cells of these animals contain numerous small granules (type A and B) and also large masses of secretory material (type C secretory material) in the basal and, in some specimens, also in the supranuclear regions of the cell (stage 2-3). Group B: Adrenalectomized animals. 18 days after adrenalectomy, cells of the SCO are in stage 1; this situation persists in specimens sacrificed up to 70 days after operation (Fig. 1). Group C: Adrenalectomy + room temperature 28 ~C. After 5 days of treatment the cells show stage 3 of secretory activity; large masses of secretory material appear in the supranuclear region. After 18 days, the secretory activity gradually increases and the cells are completely filled with secretory material (stage 4). After 30 days, there is a decrease in the secretory activity (Fig. 1). GroupD: Castrated + adrenalectomized animals. 15 days after the operation the cells of the SCO contain only small secretory granules located in the apical region

Mar. c a s t r a t i o n + ad r e n a l e c t o m y

Mar. c a s t r a t i o n + a d r e n a l e c t o m y + 28~

30

I,

1

Mar. c a s t r a t i o n + adrenatect. + testost.

1

~5 Mar. a d r e n a l e c t o m y 18

Mar. adrenalect. + 28~

30

11

May. cyproteron + 28 ~

18

I

30

1 57

7O

Stage 1

79

&5

[ ] Stage 2

~

Stage 3

72

9 Stage 4

Fig. 1. Activity stages of the subcommissural organ under experimental conditions. The numbers indicate days of treatment. For histological details, see D'Uva et al. (1977) M ar March; M a y May

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(stage 1); this condition is present in animals killed up to 60 days after the surgical treatment (Fig. 1). Group E: Castration + adrenalectomy + room temperature 28 ~C. Cells of the

SCO remain at stage 3 of the secretory cycle (Fig. 1). Their secretion is characterized by small granules and large masses of secretory material in the basal and in the supranuclear portions of the cells. During this treatment, however, the death rate is very high (about 80 ~). Group F: Castration + adrenalectomy + testosterone. 17 days after the operation

the SCO cells contain only a few small granules in apical location; the basal portion of the cell is almost free of secretory material. The activity of these cells corresponds to stage 1. Group G: Cyproterone + room temperature 28~

Treatment with cyproterone and increase in room temperature to 28 ~C in May, when the secretory activity of the SCO is very high, results in a histological picture similar to that obtained after castration or with cyproterone treatment at lower temperatures; it indicates an inhibition of the SCO. 65 days after the beginning of the treatment, the secretory activity is reduced and the cells are in stage 2 of the secretory cycle. The present observations on the SCO of Lacerta s. sicula Raf. indicate that adrenalectomy inhibits the activity of the SCO which remains at stage 1. Normal animals investigated in May show stage 3 of the secretory activity. After adrenalectomy + castration, the decrease in the secretory activity of the SCO is more rapid; about two weeks after operation there is a conspicuous decrease in the secretory activity of the SCO cells and, in May, a condition similar to that observed during the winter months is produced. In contrast, an increase in the ambient temperature in adrenalectomized animals induces a recrudescence of the activity of the SCO which reaches its maximum approximately after two weeks of treatment (stage 4). The increase in temperature in castrated + adrenalectomized lizards does not affect the inhibition produced by this type of surgical treatment; however, the elevation of the temperature is followed by a very high death rate. The administration of testosterone does not induce an initiation of secretory activity in the SCO of adrenalectomized + castrated animals, but prevents an increase in the death rate. The increase in temperature does not induce renewed activity of the SCO blocked by cyproterone administration. These experiments speak in favor of the assumption that the activity of the adrenal gland is related to the activity of the SCO. After adrenalectomy, the SCO cells of the lizard show a decrease in secretory activity; corticosteroids alone are capable of maintaining the activity of the SCO (Varano et al., 1978). The effects of castration and the response of the SCO to treatment with cyproterone that blocks the action of testosterone at the level of the peripheral target organ, seem to confirm the hypothesis that the secretory activity of the SCO parallels the sexual cycle of Lacerta s. sicula (D'Uva et al., 1977). The increase in ambient temperature alone does not effect the cells of the SCO in animals treated with cyproterone, nor does it induce a recrudescence of the SCO in castrated + adrenalectomized animals.

327

Subcommissural Organ of L a c e r t a s. sicula Raf. Table 1. Animal material; experimental and sampling procedures Group

No. Animals

Season

Treatment

Dose/injection every 3 days

Days of treatment

A

50

March-May

Controls under natural conditions

-

-

B

30

March

Adrenalectomy

-

5; 11; 18; 30; 59; 7O; 85

C

5

March

Adrenalactomy+ 28~C

-

5; 11; 18; 30; 45; 51

D

20

March

Castration+ adrenalectomy

-

7; 15; 30; 45

E

15

March

Castration + adrenalectomy + 28~

-

5; 11 ; 18

F

20

March

Castration+ adrenalectomy + testosterone

100~tg

10; 17; 26; 37

G

20

May

Cyproterone + 28~

0.5mg

57; 65; 72; 79

Since it was d e m o n s t r a t e d that d u r i n g w i n t e r b o t h the activity o f the Leydig cells a n d the f u n c t i o n o f the i n t e r r e n a l cells are strongly stimulated by a n increase in temperature, it is very p r o b a b l e that the increase in the activity o f the SCO in a d r e n a l e c t o m i z e d lizards a n d in castrated a n i m a l s kept at 28~ ( D ' U v a et al., 1978) m i r r o r s primarily a n effect exerted by high t e m p e r a t u r e o n the Leydig cells a n d the interrenal cells. I n s u m m a r y , all data o b t a i n e d in the present study a n d from the preceding investigations o n L a c e r t a s . s i c u l a p o i n t o u t that b o t h h o r m o n a l a n d thermic factors are involved in the c o n t r o l o f the secretory activity o f the SCO.

References D'Uva, V., Ciarcia, G.: The subcommissural organ of the lizard L a c e r t a s. sicula Raf. Ultrastructure during the winter. Experientia, 32, 1327-1329 (1976) D'Uva, V., Ciarcia, G., Ciarletta, A.: The subcommissural organ of the lizard L a c e r t a s. sicula Raf. Ultrastructure and secretory cycle. J. Submicr. Cyt., 8 (2-3), 175-191 (1976) D'Uva, V., Ciarcia, G., Ciarletta, A., Angelini, F.: The subcommissural organ of the lizard L a c e r t a s. sicula Raf. during the sexual cycle in normal and experimental conditions. Mort. Zool. Ital. N.S., 11 (3-4), 193-210 (1977) D'Uva, V., Ciarcia, G., Ciarletta, A., Angelini, F.: The subcommissural organ o f L a c e r t a s. sicula Raf.: effects of experimental treatments on castrated animals during winter. J. Exp. Zool., 205 (2), 285-292 (1978) Varano, L., Laforgia, V., D'Uva, V., Ciarcia, G., Ciarletta, A.: Possiblerelationship between the activity of the adrenal gland and the subcommissural organ in the lizard L a c e r t a s. sicula Raf. Cell Tissue Res., 192, 53-65 (1978) Accepted April 25, 1979