Effects of bromantan on offspring maturation and development of reflexes
Iezhitsa, I. N.; Spasov, A. A.; Bugaeva, L. I.
Neurotoxicol. Teratol. (2001), 23(2), 213-222.
Abstract
Bromantan (N-[2-adamantil]-N-[para-bromphenyl]amine) is an "actoprotective" drug widely used in Russia as a muscle performance-enhancing agent for sportsmen and as an immunostimulator in medicine. Experiments were conducted to determine whether this compound has adverse effects on the reproduction and development of offspring. Sexually mature female rats, weighing 180¯200 g, were orally given bromantan at doses of 30 mg/kg (30-mg/kg group), 150 mg/kg (150-mg/kg group) and 600 mg/kg (600-mg/kg group) daily for 16 days, while the controls received the vehicle, amylaceous solution. Afterwards, treated females were mated with untreated males. The body weight change of the pregnant rats was monitored, as well as the length of gestation, litter size, sex ratio and number of stillborn. The offsprings were weighed and observed for external malformations, abnormalities of conditioned and unconditioned reflexes and open-field behaviour. Observation of rat dams revealed that their general state and activity in all groups did not differ significantly both during and after bromantan treatment. Bromantan had no adverse effects on body weight and gestation length of dams. Number of dams delivered per group did not differ from controls. There were stillborn rat pups in all litters, but the control group had less. One dam in the first group delivered a rat pup with a head hematoma. Litter size of the 30- and 600-mg/kg groups was decreased (by 34.9% and 44.2%, respectively) and increased in the 150-mg/kg group (by 45.1%, P<.05) in comparison with controls. Bromantan had insignificant different effects on the sex ratio of newborn in all treatment groups. Survival of pups over the first 3 months showed a loss of 40% for the 150-mg/kg group and 20% for controls. During the remaining time, death rate did not exceed 3¯6% and did not differ from those of the controls. Pups in the 30- and 600-mg/kg groups showed significantly higher weight gain during the first week (7th PND) of observation by 83.69% and 58.02%, respectively, compared to controls; subsequently, this difference in the 600-mg/kg group decreased rapidly to insignificant levels, but the 30-mg/kg group remained significantly different until PND 35 and then again at PND 77¯112. Dynamics of body weight gain of rat pups in the 150-mg/kg group during the whole (but not on PND 7) period of observation was insignificantly (on PNDs 14, 42 and 49 significantly) lower than that of the control group. Study of the functional state of rat pups' nervous system at different stages of postnatal development revealed insignificant differences in the expression of reflexes compared with those of the control group. Negative geotaxis was completed by the 8th day in controls and in treated groups earlier by an average of 1¯2 days. Surface righting was completed by the controls on the 8th day, in the 30-mg/kg group on the 6th day and in other treated groups on the 7th day. Cliff avoidance appeared a day ahead for rat pups in treated groups compared with controls. Air righting reflex in the 30- and 600-mg/kg groups was observed a day earlier than in the controls. Significant differences were observed only for two parameters (negative geotaxis and surface righting); in both cases, rat pups of the 30-mg/kg group differed from the control and 150-mg/kg groups. Early development of physical parameters was also noted, but significant differences from the control group were obtained only in the 30-mg/kg group for incisor eruption. While all pups demonstrated strength of fore and hind limbs by postnatal day 16, treated pups increased their times of maintaining their grasp (PND 15). Open-field testing (PND 40) resulted in an insignificant decrease of exploratory and locomotor behaviors for the 30-, 150- and 600-mg/kg groups. The number of grooming episodes was insignificantly decreased for the 30-mg/kg group and insignificantly increased for the 600-mg/kg group. In the passive avoidance testing, on the retention day (72 h later), entry latency for rat pups in the 30-, 150- and 600-mg/kg groups increased by 259.0%, 175.3% and 160.7%, respectively (P<.05), over their training day, while in the control group, time increased only by 1.8%.
1. Introduction
Bromantan (N-[2-adamantil]-N-[para-bromphenyl]amine) is an "actoprotective" drug [5, 16 and 29] widely used in Russia as a muscle performance-enhancing agent for sportsmen [26] and is also used as immunostimulator in medicine [13].
Actoprotectors (from Latin "aktus") was created at the Pharmacology Department of Medical Military Academy (St. Piterburgh) as a result of the search for medicines that are able to support motor activity and work capacity (mostly physical) under complicated conditions (oxygen deficiency, high environmental temperature, etc.) [5, 24, 29 and 34]. In the 1960s, research of actoprotectors under the supervision of Prof. V.M. Vinogradov was done to create drugs of unexhausting action that would surpass adaptogenes, psychostimulators and other known drugs in activity, especially under complicated conditions [5 and 34]. Bromantan is one of the first well-studied actoprotector drugs for the correction of the processes of work capacity rehabilitation both in ordinary and complicated conditions.
Bromantan was developed in the 1980s at the Institute of Pharmacology of the Russian Academy of Medical Sciences (Moscow) as a drug having psychoactivating and adaptogene properties under complicated conditions (hypoxia [18], high environmental temperature [4 and 24], physical overfatigue [18 and 24], emotional stress [16 and 18], etc.). Bromantan do not concede well-known psychostimulant of phenylalkylamine structure and its analogs (amphetamine, sydnicarb, meridil, etc.) by specific activity [16]. In contrast, bromantan has neither addictive potential nor reveals redundant and exhausting activation of sympaticoadrenergic system, or decelerates the restoring of work capacity at preventive application before forthcoming activity in complicated conditions (hypoxia, high environmental temperature, physical overfatigue, emotional stress, etc.) [16]. Bromantan has no prohypoxic activity [16].
The action of bromantan (20 and 50 mg/kg po) on spectra power EEG by Fourier in the sensorimotor cortex, dorsal hippocampus and lateral hypothalamus of the left and right rat brain hemispheres in free behavior has been investigated [13]. Bromantan leads to decreases in the total and absolute powers of all frequency bands of EEG spectra, changes structural spectra in the cortex and in hippocampus ¯¯ decreases in the relative power of theta-band and increases in the relative power of beta 1, 2-activity [13]. The basic feature of bromantan's action is a two-phase effect (its maximum occurs 2¯3 and 6¯7 h after administration), which remains up to 8 h of EEG recording. In comparison with other adamantane psychostimulants, maximum EEG changes occur at 1¯1.5 h and lasted up to 4¯5 h after administration adapromine, and maximum effect of midantan occurs at 1¯3 h and remained up to 5 h after its administration. These data suggest that bromantan has more prolonged stimulant properties than other adamantane psychostimulants [13]. Bromantan, in contrast with adapromine and midantan, has more expressed effect on biopotentials of rat brain, that is reflected both in more potent quantitative modification of EEG characteristics and changes of practically all characteristics of EEG power spectra [13].
Positive effects of bromantan on the physical efficiency are associated not only with its psychostimulating action but also with the membrane-protecting effect [17].
Bromantan is used as immunostimulator in the clinic [11] as well. Bromantan has been shown action at secondary immunodeficit condition (influencing the physical overfatigue, emotional stress, effect of toxic xenobiotics and bacterial infection). Bromantan activates humoral (is more expressed) and cellular parts of immune system [11].
Bromantan action is studied insufficiently. It is known that in formation of psychotropic and immunomodulated effects of bromantan, the important role belongs in the dopamine and serotonin systems. So, for example, amphetamine-induced stereotypies, which depend on stimulation of striatal D2-receptors, were agonized by bromantan (30 and 600 mg/kg po) [31, 32 and 35]. The data on the elimination of catalepsy by bromantan (5 mg/kg ip), caused by neuroleptics haloperidol (1 mg/kg ip) and trifluoperazine (2 mg/kg ip), indirectly confirm participation of dopamine and serotonin structures in pharmacological action of the drug [16 and 19].
Pharmacological and biochemical studies also indicate that bromantan exposure affects brain dopamine systems [12 and 19]. Bromantan (5¯50 mg/kg ip) has expressed central dopaminomimetic effect, connected to an amplification of dopamine release from presynaptic ending. Bromantan at 50¯500 mkM significantly decreased uptake of serotonin ¯¯ by isolated rats' cerebral synaptosomes and to a lesser degree inhibited uptake of dopamine [12, 16 and 19]. The central adrenergic effect of the drug is less expressed: blocking of the synaptosomal capture of norepinephrine in higher (more than 500 mkM) concentrations, and absence of an effect on the mediator content in rat brain [19].
Bromantan (50¯500 mkM) caused expressed and prolonged (registered during 8 h) increase of dopamine release [12]. Tetrodotoxin (10-6 M), perfused through microdialysis probe, partly antagonized the increase of dopamine expression caused by bromantan [12]. Extracellular levels of dopamine metabolites 3,4-dioxyphenilacetic and homovanillic acids decreased insignificantly [12 and 14].
In a number of works, the hypothesis that anxiolytic effect is present in the spectrum of pharmacological properties of bromantan is stated [25 and 26]. This is shown by per os administration of bromantan in doses of 40¯50 mg/kg, which prevents the fixed posture reaction and reduces the level of defecation in Balb/c mice in the "open-field" test and activates their behavior in the "elevated plus maze" test. It also increases the motor activity of C57Bl/c mice in both tests [25 and 26].
We hypothesize that bromantan, exerting a dopaminergic effect on activity [12, 19, 31 and 32], may probably depress prolactin secretion of lactating rats. There is sufficient evidence that the influence of dopaminomimetic drugs (i.e., bromocriptine) leads to suppression of prolactin in the blood of lactating rats on the 5th day of lactation [2, 15 and 20]. Prolactin is the main hormone of lactogenesis, and suppression of its secretion at the beginning of lactation influences milk secretion [3, 8 and 9]. It has further been shown to decrease body weight gain, thus, influencing onset of pubescence and physiological development [2 and 15]. Deficiency of this hormone also influences the mechanisms of sexual differentiation of cerebrum [3, 21, 22 and 23].
The early period of postnatal ontogenesis is the critical period in cerebral development and the quantity of hormones and neurotransmitters, as well as morphological maturation and development of the neuroendocrine system [3, 21, 22 and 23]. It has been shown that the decline in normal concentrations of neurotransmitters and hormones during this critical period may cause permanent changes of physiological functions, such as sexual behaviour, emotional state, immunity and ability to learn [2, 3, 15, 21, 22 and 23]. This early postnatal period coincides with lactation of mammals, and dam's milk is the source of hormones (i.e., prolactin) [2, 3 and 15].
Proceeding from above, we hypothesize that bromantan could affect on physical maturation of progeny. In light of this, the aim of this study was to observe the effects of the actoprotective (muscle performance enhancing) drug, bromantan, on the development of rat pups, whose
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