Tricarbonyl(methylcyclopentadienyl)manganese

Specific investigations: other studies

Additional information

Two in vitro and two in vivo studies providing information on the effects of Mn (iin the form of mmt) are included

In vitro

Anantharam (2002)

The oxidative stress-dependent cellular events in dopaminergic cells after exposure to a mmt In pheochromocytoma cells was characterizzed. mmt exposure resulted in rapid increase in generation of reactive oxygen species (ROS) within 5 -15 min, followed by release mitochondrial cytochrome C into cytoplasm and subsequent activation of cysteine proteases, caspase-9 (two-fold to three-fold), and caspase-3 (15- to 25- fold), but not caspase-8, in a time- and dose-dependent manner. mmt exposure induces a time- and dose-dependent porteolytic cleavage of native protein kinase Cδ (PKCδ, 72 -74 kDa) to yield 41 kDa catalytically active and 38 kDa regulatory fragments. Pretreatment with caspase inhibitors (Z-DEVD-FMK or Z-VAD-FMK) blocked mmt-induced proteolytic cleavage of PKCδ, indicating that cleavage is mediated by caspase-3. Furthermore, inhibition of PKCδ activity with a specific inhibitor, rottlerin, significantly inhibited caspase-3 activation in a dose-dependent manner along with a reduction in PKCδ cleavage products, indicating a possible positive feedback activation of caspase-3 activity by PKCδ. The presence of such a positive feedback loop was also confirmed by delivering the catalytically active PKCδ fragment. Attenuation of ROS generation, caspase-3 activation, and PKCδ activity before mmt treatment almost completely suppressed DNA fragmentation. Additionally, overexpression of catalytically inactive PKCδ^K376R (dominant negative mutant) prevented mmt-induced apoptosis in immortalized mesencephalic dopaminergic cells. These data demonstrate that caspase-3 -dependent proteolytic activation of PKCδ plays a key role in oxidative stress-mediated apoptosis in dopaminergic cells after exposure to mmt.

Kitazawa (2002)

mmt was investigated to determine whether it potentially causes dopaminergic neurotoxic effects. mmt is acutely cytotoxic with dopamine-producing cells (PC-12) seemed to be more susceptible to cytotoxic effects than nondopaminergic cells (striatal γ-aminobutyric acidergic and cerebellar granule cells). mmt also potently depleted dopamine apparently by cytoplasmic vesicular release to the cytosol, a neurochemical change resembling other dopaminergic neutoxicants. Generation of reactive oxygen species (ROS), an early effect in toxicant-induced apoptosis, occurred within 15 minutes of mmt-exposure. mmt caused a loss of mitochondrial transmembrane potential (∆ψm), a likely source of ROS generation. The ROS signal further activated caspase-3, an important effector caspase, which could be inhibited by antioxidants (Trolox or N-acetylcysteine). Predepletion, of dopamine by using α-methyl-ρ-tyrosine (tyrosine hydroxylase inhibitor) treatment partially prevented caspase-3 activation, denoting a significant dopamine and/or dopamine by-product contribution to initiation of apoptosis. Genomic DNA fragmentation, a terminal hallmark of apoptosis, was induced concentration dependently by mmt but completely prevented by pretreatment with Trolox, deprenyl (monoamine oxidase-B inhibitor) and α-methyl-ρ-tyrosine. A final set of critical experiments was performed to verify the pharmacological studies using a stable Bcl-2-overexpressing PC-12 cell line. Bcl-2-overexpressing cells were significantly refractory to mmt-induced ROS generation, caspase-3 activation, and loss of ∆ψm and were completely resistant to MMT-induced DNA fragmentation. Taken together, the results presented herein demonstrate that oxidative stress plays an important role in mitochondrial-mediated apoptotic cell death in cultured dopamine-producing cells after exposure to mmt.

In vivo

Liu (1999)

The in vivo effect of mmt was studied on the motor nerve of the mouse. Adult mice of the ICR strain, weighing 25 -30g were pretreated intraperitoneally with mmt in corn oil (0.05 and 0.01 mg/g/day for 3 days). The motor nerve conduction velocity was markedly decreased in mmt-treated mice. The Na+, K+-ATPase activity of sciatic nerve isolated from mmt-treated mice was decreased; however, the sciatic nerve Na+, K+-ATPase activity was not affected by the in vitro treatment of mmt. Moreover, [3H]ouabain binding of sciatic nerve isolated from mmt-treated mice was decreased. Using Western blot analysis, the amount of Na+, K+-ATPase catalytic α1 subunit polypeptide in sciatic nerve of mmt-treated mice was also decreased. These results indicate that a causal relationship may exist between reduced nerve Na+, K+-ATPase activity and motor nerve conduction velocity in mmt-treated mice and that a measurable decrease in α1 catalytic subunit isoform of Na+, K+-ATPase may be necessary for the development of peripheral neuropathy by mmt.

Gianatsus (1985)

Mice received an acute sc dose of manganese chloride, oxide (Mn3O4) or mmt (0.2 or 0.4meq/kg). A single injection markedly elevated brain manganese concentrations within 1 day with elevated levels maintained for at least 21 days. Repeated injections led to further increases in both brain and blood, although the levels in the brain appeared to persist at consistently high levels for longer periods. The chloride form produced higher brain levels than either of the other two forms. These results appear to suggest that the slowly developing neurotoxicity in response to manganese exposure may be due to a persistent retention of manganese by the brain.  The oxide was chosen because is the combustion product of mmt, while the dichloride form is a water soluble salt of the metal.

Both brain and blood manganese levels were elevated after injection, but there was a slight difference in the time course of this effect between the two tissues. There was also a difference dependent on the form of Mn tested. With the oxide form, blood manganese reached a maximum 4h after injection and remained elevated after 1 week. In the brain, Mn reached is peak at 24h after administration, remained also constant for 1 week. With the dichloride form, peak blood levels reached extremely high values (more than 10 000% of control) within 1h and gradually declined over the course of 1 week (although they were still 740% of control at 1 week). In the brain peak levels occurred approximately 24h after injection and remained at this level for at least 1 week.

A similar trend was observed with repeated injections. Higher brain and blood levels of Mn were detected with repeated injections of either compound than following an acute injection. However, the elevated brain Mn levels appeared to be less readily reversible than the blood; brain levels 21 days after a single injection of either form of Mn were no different from the levels observed 7 days after a single injection. Blood levels, although still significantly elevated above the control by 21 days, had significantly declined compared with values obtained 1 week after injection. Thus, the exit of Mn from the brain, appears to be a slower process than the entry of the metal.