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This study is aimed at exploring the possible mechanism of hypnosis-enhancing effect of HgS or cinnabar (a traditional Chinese medicine containing more than 95% HgS) in mice treated with pentobarbital. We also examined whether the effect of HgS is different from that of the well-known methyl mercury (MeHg). After a short period (7 days) of oral administration to mice, a nontoxic dose (0.1 g/kg) of HgS not only significantly enhanced pentobarbital-induced hypnosis but also attenuated tolerance induction; while a higher dose (1 g/kg) of HgS or cinnabar exerted an almost irreversible enhancing effect on pentobarbital-hypnosis similar to that of MeHg (2 mg/kg) tested, which was still effective even after 10 or 35 days cessation of administration. To study comparatively the effects of different mercury forms from oral administration of MeHg and HgS on membrane ATPase activities of experimental mice, analysis of the Hg content in the cerebral cortex revealed that correlated with the decrease of Na(+)/K(+)-ATPase and Ca(2+)-ATPase activities. Furthermore, NO levels of blood but not that of cerebral cortex were also decreased by mercuric compounds. Although pentobarbital alone enhanced cytochrome p450-2C9 in time dependent manner, all of mercurial compounds tested had no such effect. All of these findings indicated that the mercurial compounds including cinnabar, HgS and MeHg exert a long-lasting enhancing hypnotic activity without affecting pentobarbital metabolism, which provides evidence-based sedative effect of cinnabar used in Chinese traditional medicine for more than 2,000 years. The nontoxic HgS dosing (0.1 g/kg/day) for consecutive 7 days is perhaps useful for delaying or preventing pentobarbital-tolerance.

Methylmercury (MeHg) is an environmental toxicant, while mercuric sulfide (HgS) is a main active component of cinnabar, a Chinese mineral medicine used as a sedative. Because the neurotoxicological effects of HgS were not clearly understood, in this study, we attempted to compare HgS with MeHg in various physiological responses in Sprague-Dawley rats. After oral administration (2 mg/(kg day)) for consecutive 5 and 14 days, MeHg reversibly decreased both of motor nerve conduction velocity (MNCV) and tail flick response, whereas irreversibly inhibited all of the motor equilibrium performance, recovery of compound muscle action potentials (CMAP) following exhaustic tetanic stimuli and Na+/K+-ATPase activity of the isolated sciatic nerve. These toxic effects of MeHg were found in well correlation of Hg contents of various tissues (blood, cerebral cortex, liver and kidney) in rats. For comparison, a dose of 1g/(kg day) of HgS was orally administered to the rats based on our previous findings on ototoxicity of HgS. The results revealed that HgS only reversibly delayed the recovery of suppressed CMAP and inhibited sciatic nerve Na+/K+-ATPase activity in accordance to the lower Hg contents of the tissues. These findings provide the important information on the differential susceptibility of various nervous tissues to MeHg and HgS. The neruotoxic effects produced by HgS was estimated to be about 1000 of those induced by MeHg found in this study and our previous reports.

Cinnabar, a naturally occurring mercuric sulfide (HgS), has been combined with Chinese herbal medicine as a sedative for more than 2000 years. To date, its neurotoxic effect on the vestibulo-ocular reflex (VOR) system has not been reported. By means of a caloric test coupled with electronystagmographic recordings, the effect of commercial HgS and cinnabar on the VOR system of guinea pigs was studied. HgS or cinnabar was administered orally (1.0 g/kg) to Hartley-strain guinea pigs once daily for 7 consecutive days. A battery of electrophysiological, biochemical, and histopathological examinations were performed. The results showed that HgS induced a 60% caloric response abnormality (40% caloric hyperfunction and 20% hypofunction), whereas the abnormal responses appeared to be more severe (six out of six) in the cinnabar group. The Hg contents of whole blood and cerebellum were increased and correlated to their neurotoxic effects on the VOR system, indicating that both insoluble HgS and cinnabar could be absorbed from the gastrointestinal tract and distributed to the cerebellum. Although the vestibular labyrinth revealed no remarkable change under light microscopy, loss of Purkinje cells in the cerebellum was detected, and the enzymatic Na(+)/K(+)-ATPase activity of cerebellum (a higher inhibitory center of the VOR system) was significantly inhibited by HgS and cinnabar. Moreover, cerebellar nitric oxide (NO) production was increased significantly. Hence, we tentatively conclude that the increased Hg contents in the cerebellum following oral administration of HgS and cinnabar were responsible, at least in part, for the detrimental neurotoxic effect on the VOR system. Potentially, decreasing Na(+)/K(+)-ATPase activity and increasing NO production within the cerebellar regulatory center are postulated to mediate this VOR dysfunction caused by the mercurial compounds and cinnabar.

A traditional Chinese mineral medicine, cinnabar, naturally occurring mercuric sulfide (HgS), is still occasionally prescribed, but the neurotoxic effects of HgS have not been elucidated. In this paper, an animal model of the purified HgS intoxication was established in guinea pigs in order to study neurotoxicity and pathophysiology of the vestibular ocular reflex system (VOR). Guinea pigs were dosed with HgS by gastric gavage (0.01, 0.1 and 1.0 g/kg per day) for 7 consecutive days. By means of caloric testing coupled with the electronystagmographic (ENG) recording in guinea pigs, we have found that HgS at a dose of 0.1 g/kg induced reversible caloric hypofunction pattern and at a higher dose of 1.0 g/kg induced irreversible hypofunction of caloric test. Monitoring the mercury contents of various tissues (blood, kidney, liver and cerebellum) by continuous flow and cold vapor atomic absorption spectrometry (AAS) revealed that a certain amount of HgS could be absorbed from the gastrointestinal tract and was detectable in these tissues. In addition to the induced dysfunction of VOR system, HgS also caused disturbance of motor performance in guinea pigs. In enzyme assay, Na+/K+-ATPase activity of cerebellum was also significantly inhibited by HgS. Morphological studies showed partial cell loss only in the cerebellar Purkinje cell layer, but not in the granule cell layer, nor in the vestibular labyrinth. All of these findings suggest that cerebellar Purkinje cells are the sensitive target site responsible for HgS-inducing dysfunctions of both VOR system and the motor performance in guinea pigs. Thus, it is concluded that caloric test coupled with ENG recording in VOR system is certainly a sensitive biomarker for monitoring the neurotoxicity of HgS.

In this paper, we attempted to construct an animal (mouse) model for monitoring the oto-neurotoxicity of mercuric sulfide, comparing its toxicity with the well-known (organic) mercury compound methyl-mercury. Mice were treated with either mercuric sulfide (HgS, 0.1 and 1.0 g/kg per day) or methyl-mercury (MeHg, 0.2, 2.0 and 10 mg/kg per day) by gastric gavage for 7 consecutive days. Analysis of auditory brainstem response (ABR) indicated that significant elevation of the physiological hearing threshold as well as significant prolongation of interwave latency I-V was observed for MeHg -- (2.0 and 0.2 mg/kg per day) or HgS -- (1.0 g/kg per day, but not 0.1 g/kg per day) treated mice. Further, both MeHg- and HgS-treated animals demonstrated a significant prolongation of interwave latency I-V that increased with an increasing mean blood-Hg level. The oto-neurotoxicity of MeHg (2.0 mg/kg per day) persisted to at least 11 weeks subsequent to the cessation of its administration. The toxic effect of HgS, however, disappeared completely 5 weeks subsequent to the cessation of its administration. These results suggest a correlation between the Hg-elicited hearing dysfunction and the availability of mercury in brain tissue. Both inhibition of Na(+)/K(+)-ATPase activity and overproduction of nitric oxide in the brainstem are consistent with an analysis of the physiological hearing threshold and latencies of ABR waveform at all time points throughout the experimental process. Thus, it is proposed that high-dose HgS or MeHg intoxication is associated with a decrease in functional Na(+)/K(+)-ATPase activity in the brainstem of affected animals, this presumably arising via excessive nitric oxide production, and suggesting that brainstem damage may play a role in mercury-induced hearing loss.

This study compared the neurobehavioral toxicities of three mercurial compounds: methyl mercury (MeHg) which is soluble and organic. and mercuric sulfide (HgS) and cinnabar (naturally occurring HgS), which are insoluble and inorganic. Cinnabar, a Chinese mineral medicine, is still used as a sedative in some Asian countries, but there is relatively little toxicological information about it. These mercurial compounds were administered intraperitoneally (MeHg, 2 mg/ kg) or orally (HgS and cinnabar, 1.0 g/kg) to male rats once every day for 13 consecutive days with assays conducted during or after discontinuous administration for 1 h, 2, 8 and 33 weeks. Neurotoxicity was assessed based on the active avoid-ance response and locomotor activity. The results obtained showed that MeHg and cinnabar prominently and irreversibly caused a decrease in body weight, prolongation of latency for escape from electric shock, a decrease in the percentage for the conditioned avoidance response (CAR) to electric shock, impairment of spontaneous locomotion and inhibition of Na+/K+-ATPase activity of the cerebral cortex. In contrast. HgS reversibly inhibited spontaneous locomotion and Na+/K+-ATPase activity. It was noted that HgS significantly decreased the latency of escape from electric shock during the ad-ministration period, which lasted for 33 weeks after discontinuous administration. In fact that pretreatment with arecoline (a cholinergic receptor agonist) but not fipexide (a dopaminergic receptor agonist) could significantly shorten the prolonged latency for escape caused by MeHg and cinnabar, suggested that the deficit in the active avoidance response was perhaps, at least in part, mediated by the dysfunction of the cholinergic rather than the dopaminergic system. Determination of the Hg levels of the whole blood and cerebral cortex revealed that the tissue mercury content was highly correlated with the degree of neurobehavioral toxicity of these Hg compounds. These findings suggest that insoluble HgS and cinnabar can be absorbed from the G-I tract and distributed to the brain. The possibility that contamination due to other minerals in the cinnabar is responsible for the greater neurotoxic effects compared to HgS is under investigation.