Effects of REM deprivation on the lordosis response induced by gonadal steroids in ovariectomized rats

Physiology & Behavior, Vol. 32, pp. 91-94. Copyright©PergamonPress Ltd., 1984. Printedin the U.S.A.

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Effects of REM Deprivation on the Lordosis Response Induced by Gonadal Steroids in Ovariectomized Rats J. VELAZQUEZ-MOCTEZUMA, E. MONROY, C. B E Y E R A N D E. C A N C H O L A

D e p a r t a m e n t o de Biologia de la Reproduccion, Universidad A u t o n o m a Metropolitana Iztapalapa A . P . 55535, Mexico City, C.P. 09348, Mexico R e c e i v e d 21 M a r c h 1983 VELAZQUEZ-MOCTEZUMA, J., E. MONROY, C. BEYER AND E. CANCHOLA. Effects of REM deprivation on the lordosis response induced by gonadal steroids in ovariectomized rats. PHYSIOL BEHAV 32(1) 91-94, 1984.-Ovariectomized rats were submitted to REM sleep deprivation (REMd) using the water tank technique and their behavioral responsiveness (lordosis) to gonadal steroids was tested. In Experiment 1, animals received 2/xg of estradiol benzoate (EB) followed by 2 mg of progesterone (P) 44 hours later. Several REMd periods (12, 72, 96 and 120 hr) were applied, all ending four hr after P. REMd animals showed significantly lower lordosis quotients (LQ) than undisturbed sleep animals regardless of the duration of the deprivation period. In Experiment 2, animals received a single dose of EB (2/zg) and were REMd for 120 hours. Animals were tested daily to evaluate their LQ. EB, at this dose level, failed to elicit significant lordosis behavior in undisturbed sleep rats. REMd rats gradually increased their LQ values reaching maximal levels at 72 hours. Adrenalectomized control groups receiving the same hormonal treatment responded similarly to the experimental groups, thus discarding the participation of adrenal steroids in these effects. The present results show that REMd differentially affects the response to E and P in ovariectomized rats, enhancing the former and inhibiting the latter. REM sleep deprivation

Estrogen

Progesterone

Sexual behavior

Group I. Control, 10 rats, EB 2/xg plus 2 mg P, undisturbed sleep cycle. Group 2. Fifteen rats, EB 2/xg plus 2 mg P, REM sleep deprivation for 12 hr. Group 3. Sixteen rats, EB 2/xg plus 2 mg P, REM sleep deprivation for 72 hr. Group 4. Fifteen rats, EB 2/xg plus 2 mg P. REM sleep deprivation for 96 hr. Group 5. Sixteen rats, EB 2 p.g plus 2 mg P, REM sleep deprivation for 120 hr. EB was injected subcutaneously in a volume of 0.1 ml of sesame oil at time 0. P was injected subcutaneously in the same volume 44 hours after estrogen. REM sleep deprivation was achieved by placing the rat on a 6.5 cm platform surrounded by water up to 1 cm beneath the surface [7,12]. All REM deprivation periods ended 4 hours after P administration. Ss were tested half an hour before P injection (43:30 hr) and at 48,52, 72, 96 and 120 hr after EB administration.

REM sleep deprivation induces alterations in several behavioral responses (for review see [2, 9, 15]). Interestingly, some of these behaviors, such as aggressiveness, food intake and locomotor activity have been reported to be under steroid hormone control. Moreover, it has also been reported that REM sleep deprivation diminishes the effects of estradiol benzoate on feeding and locomotion [8]. Nevertheless, to our knowledge there is only one report concerning the relationship between REM sleep and reproductive behavior [16]. Therefore, in the present study we analized the effect of REM sleep deprivation on the sexual responsiveness of ovariectomized rats to either estrogen or estrogen plus progesterone treatments. METHOD One hundred-twelve Wistar rats weighing 250-300 g were used in this study. The subjects (Ss) were housed in large cages (6 per cage) and maintained in a room with reversed light cycle (lights on 2000: off 1000). Food (Purina Chow) and water were available ad lib. Ss were ovariectomized under ether anesthesia at least two weeks before hormonal treatment.

Experiment 2 The effect of REM deprivation on the lordotic response to 2 txg of EB was studied in the following three groups: Group 6. Ten rats, REM sleep deprivation for 120 hours. The initiation of REM deprivation coincided with the injection of sesame oil. Group 7. Ten rats, EB 2/xg, undisturbed sleep cycle. Group 8. Ten rats, EB 2/zg, REM sleep deprivation for

Experiment I The effect of REM deprivation on the lordotic response to the sequential administration of estradiol benzoate (EB) and progesterone (P) was studied in the following five groups:

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FIG. 1. Effect of different periods of REM sleep deprivation on the lordosis response induced by progesterone in estrogen-primed rats. Note clear decrease in responsivity to progesterone in REM deprived rats (groups 2 to 5) in comparison to control group (group 1).

TABLE 1 SIGNIFICANTp VALUES* AMONG GROUPS IN EXPERIMENT I Control Group vs. Experimental Groups Groups Control vs. 2 Control vs. 3

Control vs. 4 Control vs. 5

Among Experimental Groups

p Values p<0.001 p<0.001 p<0.001 p<0.001 p<0.050 p<0.001 p<0.001 p<0.025 p<0.010

at 48 hr at 52 hr at 48 hr at 52 hr at 72 hr at 48 hr at 52 hr at 48 hr at 52 hr

Groups

p Values

2 vs. 3

p<0.01 at 52 hr

2 vs. 4

p <0.025 at 44 hr p<0.050 at 52 hr

*Mann Whitney's U test. See text for explanation.

120 hours. Initiation of REM deprivation coincided with EB injection. In order to analyze the possible participation of adrenal steroids in changes induced by REM deprivation, the following three groups were studied: Group 9. Nine rats, adrenalectomized, REM sleep deprivation for 120 hours, REM sleep deprivation coincided with sesame oil injection. Group 10. Nine rats, adrenalectomized, EB 2 ttg plus 2 mg P, REM sleep deprivation for 48 hours. Initiation of REM sleep deprivation coincided with EB injection and P was administered 44 hours later. Group 11. Nine rats, adrenalectomized, EB 2 ttg, REM sleep deprivation for 120 hours. Initiation of REM sleep deprivation coincided with EB injection. Sesame oil injection at 44 hours.

In groups 6-11, Ss were tested at 0, 24, 44, 48, 52, 72, 96 and 120 hours after EB administration. Instead of testing group 10 at 44 hours, it was tested at 43:30 hours. Animals were adrenalectomized 10 days before the initiation of the experiment. In adrenalectomized rats, drinking water was substituted with a 0.9% saline solution. Tests were made in a circular observation cage (Plexiglas) of 53 cm in diameter. Sires were vigorous, experimented males. Each female received 10 vigorous mounts in every test. Receptivity of the females was quantified by a lordosis quotient ( L Q = n u m b e r of lordosis/10 × 100). Statistical analysis was made using the Mann Whitney U test. RESULTS Figure 1 summarizes the results obtained in Experiment 1. It can be seen that experimental groups (REM deprived

REM DEPRIVATION AND SEXUAL BEHAVIOR

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FIG. 2. Effect of an 120 hour period of REM sleep deprivation on the lordosis response induced by a single injection of 2/~g of estradiol benzoate in ovariectomized rats. Note clear facilitation of the lordosis response (time 0) to estrogen administration.

, % Of receptive subjects ~ DeDrivatK:.~nperod

FIG. 3. Changes in lordosis behavior induced by REM sleep deprivation in ovariectomized, adrenalectomized rats in response to: (a) EB 2 /xg plus 2 mg P; (b) EB treatment (2 tzg), Note that REM deprivation induced similar responses in ovariectomizedadrenalectomized rats to those in ovariectomized rats.

TABLE 2 SIGNIFICANT p VALUES* AMONG GROUPS IN EXPERIMENT 2 Groups 6 vs. 7 6 vs. 8

7 vs. 8

p Values

Hours

Non-significant p<0.025 p <0.025 p<0.010 p
All 48 52 72 96 120 72 96 120

rats) showed significantly l o w e r L Q ' s in response to P than control rats (sleep undisturbed rats). Differences b e t w e e n control and R E M deprived rats were maximal at 4 and 8 hours after P (see Table 1 for statistical comparisons). Slight non-significant differences in response to P w e r e o b s e r v e d among the experimental groups suggesting that 12 hours of R E M deprivation already produced a maximal inhibitory effect. The response to E B alone (testing time 43:30 hr) was s o m e w h a t higher in R E M deprived than in control animals. Figure 2 summarizes the results obtained in E x p e r i m e n t 2. It can be seen that in group 8 (EB plus R E M deprivation) there was a clear response to E B that gradually increased as

*Mann Whitney's U test. See text for explanation. TABLE

Significant p Values* Among Adrenalectomized Groups and Experimental Groups

Significant p Values* in Adrenalectomized Groups

Groups

p Values

Groups

p Values

5 vs. 10

Non-significant at all hr

9 vs. 10

8 vs. 11

Non-significant at all hr

Non-significant at 44 hr p<0.01 at 48 hr p<0.01 at 52 hr p<0.001 at 72 hr p<0.001 at 96 hr p<0.012 at 120 hr Non-significant at 24 hr p<0.001 at 44 hr p<0.001 at 48 hr p<0.001 at 52 hr p<0.001 at 72 hr p<0.001 at 96 hr p<0.050 at 120 hr

9 vs. 11

*Mann Whitney's U test. See text for explanation.

94

V E L A Z Q U E Z - M O ( 7 1 E Z U M A t:1 A t

REM deprivation progressed, reaching a maximal value at 72 hr. Henceforth, this high level of response was maintained throughout the observation (up to 120 hr). REM deprived rats without hormonal treatment (sesame oil injection) did not show lordosis behavior. Rats receiving EB with undisturbed sleep cycle showed only a weak behavioral response. Table 2 shows the statistical analysis of this experiment. Results obtained with adrenalectomized rats can be seen in Fig. 3. Adrenalectomized rats submitted only to REM sleep deprivation did not show lordosis behavior, while groups 10 and I I which received hormonal treatment, showed a response similar in magnitude to those observed in the ovariectomized rats (Table 3). It must be pointed out that REM deprivation in adrenalectomized rats was very deleterious, therefore the size of the adrenalectomized groups decreased as the experimental procedure progressed. Thus in group I0, N =9 at the beginning of the experiment (0-52 hr), decreased to 8 at 72 and 96 hr and to 6 at 120 hr, in group 11, N = 9 at 0 hr, 7 at 24-52 hr, 6 at 72-96 hr and is down to 3 at 120 hr. DISCUSSION Our results show that REM sleep deprivation differentially altered the response of the lordosis brain substrate to steroid hormones. Thus, REM sleep deprivation facilitated the response to estrogen alone while clearly interfered with the facilitatory effect normally exerted by P. Moreover, while the magnitude of the inhibition exerted by REM deprivation on the action of P was independent of its duration, the factlitatory effect of REM deprivation on the action ot EB alone tended to increase with the length of REM deprivation. This suggests that two different cellular mechanisms of action were influenced by REM deprivation. As REM sleep deprivation is a well established stressor, steroids of adrenal origin could be the cause of the observed phenomena. However, the results obtained in the adrenalectomized groups do not support this possibility. It has been suggested that REM sleep deprivation induces a marked decrease in the availability of catecholamines at synaptic level [14]. On the other hand, there is some evidence

that P may exert its effect through the activation of the noradrenergic system. Thus, P increases the turnover ~t' noradrenaline in the cerebral cortex and hypothalamus 16j and promotes the release of noradrenaline at hypothalamic level 110]. It has been also reported thai noradrenalinc facilitated lordosis behavior when placed in midbrain sites sensitive to P [41. Moreover, an intense lordotic response can be elicited by using amphetamine (a catecholamine releasing agent) and pimozide (a dopamine receptor blocker~ suggesting that this effect is due to a noradrenaline-receptor interaction 16]. Thus, it is possible that the inhibitory effect of REM sleep deprivation on the EB plus P action was due to an impairment in the functioning of the noradrenergic system through which P may exert its effect on the female lordosis response. Another possibility is that REM sleep deprivation interfered with P action through the inhibition of the protein synthesis and/or phosphorylation processes. It has been reported that during sleep protein phosphorylation increases two to three fold [13]. This finding primarily occurs during REM sleep 15]. On the other hand, it has been postulated that P exerts its effects on female sexual behavior by activating, through phosphorylation, the proteins previously svnthetized by estrogen [3], Therefore, it is possible that REM sleep deprivation interfered with the facilitatory effect of P on female sexual behavior by decreasing synthesis and/or reducing the phosphorylation process for activation. The increased sensitivity to estrogen caused by REM sleep deprivation is similar to that produced by lesion of the septal region 11 1], suggesting that this effecl is due lo the remotion of a tonic inhibitory influence on lordotic behavior. Moreover, it is well known that alpha-methyl-paratyrosine (a drug which interferes catecholamine synthesis) elicits a marked lordotic response in estrogen primed rats [1 I- Thus, it is possible that REM sleep deprivation removes a catecholamine-mediated inhibitory influence on estrogen action. Further research however, is required to elucidate the mechanisms through which REM sleep deprivation modulates the action of gonadal steroids on female sexual behavior.

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