6.1. Amphetamines as monoamine oxidase inhibitors
A means by which amphetamines can influence neuro- transmitter release apart from effects on plasma membrane and vesicular transporters is to increase the quantity of biogenic amine available for release by inhibiting MAOs — key mediators of amine catabolism located on the outer mitochondrial membrane. AMPH derivatives are competitive inhibitors of these enzymes, and recognition of this fact far predates the isolation of the enzymes. Hermann Blaschko et al. (1937) used a range of substrates and inhibitors to characterize amine oxidizing activities in various tissue, animal, and plant preparations. They demonstrated that many isopropylamines, including AMPH and ephedrine, are not themselves subject to degradation by the enzyme preparations but can nevertheless block the consumption of oxygen otherwise used to oxidize substrates including epinephrine and tyramine. It is worth noting Merton Sandler’s opinion that these studies of amine oxidation by Blaschko ‘‘were probably responsible for triggering off the psychopharmacology revolution’’ (Sandler, 2004) as they presaged the introduction of MAO inhibitors for treatment of depression: such use of the drugs was initiated in the early 1950s after mood elevation was noted following administration of iproniazid to tuberculosis patients.
Mann and Quastel (1940) reported an analogous finding on MAO inhibition motivated by an entirely different rationale. AMPH has long been used for treating narcolepsy (Section 2.3). The authors were interested in testing a hypothesis that narcolepsy resulted from amine-induced inhibition of brain metabolism by determining whether AMPH prevented a gradual decline in respiration observed in tissue treated with amine oxidase substrates such as tyramine. Their findings suggested that aldehydes formed by amine oxidation depressed respiration and that AMPH inhibited their formation, albeit with modest potency.
In 1971, Leitz and Stefano published a study of norepinephrine metabolism and release in which they drew the bold conclusion that the principal actions of AMPH, tyramine, and metaraminol on neurotransmitter release is mediated through MAO inhibition. Using perfused rat hearts preloaded with [3H]norepinephrine, they observed that AMPH and tyramine increased [3H]norepinephrine and concomitantly decreased its tritiated deaminated metabo- lites in the perfusate. To bolster their argument, the authors also showed that tranylcypromine (18), an irreversible MAO inhibitor which is an AMPH analog, had effects comparable to those of AMPH on releasing tritiated species. These results, however, were likely the conflated outcome of at least three distinct mechanisms: (1) effects on MAO activity; (2) efflux through plasma membrane NET; and (3) the efflux of the acidic and glycol metabolites. Furthermore, tranylcypromine is a substrate of the plasma membrane transporters and therefore could promote efflux, while the timescale tested may have been too short to observe irreversible effects of tranylcypromine on MAO. Rutledge (1970) made a concerted attempt to disentangle such factors using rabbit brain slices and synaptosomal preparations and reached a different conclusion: that AMPH prevents deamination most potently by acting at plasma membrane transporters to block reuptake and facilitate efflux of amines from the cytoplasm which thus escape MAO, although AMPH additionally acts to directly inhibit MAO.
The development of relatively specific irreversible inhibitors for MAO A (clorgyline) and B (deprenyl) (13) subtypes in the 1960s (Knoll et al., 1965; Johnston, 1968; Knoll and Magyar, 1972) facilitated the study of inhibitor selectivity. AMPH and many of its analogs show 5-fold or greater selectivity for MAO A over MAO B (Mantle et al., 1976; Robinson, 1985; Scorza et al., 1997), and most behave as reversible and competitive non-substrate inhibitors. The affinity of AMPH for MAO A is typically in the range of 10 mM for the S(+)-enantiomer, a few fold higher in affinity than its optical antipode (Mantle et al., 1976; Robinson, 1985). Thus, the stereoselectivity of MAO A for S(+)-AMPH is similar to those of DAT and VMAT. METH, which shows comparable affinity and stereoselectivity to AMPH at the plasma membrane transporters, in contrast, displays about five-fold lower affinity (rv100 mM) than (±)-AMPH for MAO A and essentially no stereoselectivity (Robinson, 1985). The affinities of AMPH and METH for MAOs compare unfavorably with their potency at NET and DAT, which are usually reported in the 100–200 nM range (references in Sections 5.1 and 5.2).
Considering the poor affinity and stereoselectivity of MAO B for AMPH and METH, it is curious that the prototypic substrate for MAO B is b-phenethylamine, which differs from AMPH only in lacking the a-methyl group, and that the first and still widely-used irreversible selective MAO B inhibitor is (-)-deprenyl, an N-2-propynyl analog of METH. [Note that deprenyl is metabolized to AMPH and METH (Reynolds et al., 1978).] AMPH-like structures have served as a useful scaffold for the development of much more potent competitive and irreversible inhibitors includ- ing amiflamine (Ask et al., 1989) (19), 4-methoxy-AMPH (PMA) (20), 4-methylthio-AMPH (Scorza et al., 1997) (21), pheniprazine (22), and tranylcypromine (18).
In summary, while there is a relatively low potency of AMPH at MAO, one cannot discount a contribution from MAO inhibition to its spectrum of physiological activity. Notably, while neither reserpine nor MAO inhibitors alone increase cytosolic dopamine, intracellular patch electro- chemistry shows that the combination of the two causes a profound increase similar to that of AMPH itself (Mosharov et al., 2003). As AMPH is a substrate of the plasma membrane transporters, it could be concentrated in the cytoplasm to a level that inhibits MAOs on the outer mitochondrial membranes. Numerous factors already mentioned make it difficult to determine the cytoplasmic AMPH concentrations attained after physiological doses of the drug, including the ability of AMPH to lipophilically diffuse across membranes and its uptake into vesicles by VMAT and due to its accumulation as a weak base in acidic compartments. Finally, although little data is available, a major metabolite of AMPH (Cho and Kumagai, 1994) 4-OH-AMPH (a- methyl-p-tyramine) (23) is likely to serve as a competitive inhibitor of MAO A. As a substrate of the plasma membrane transporters (Cho et al., 1975, 1977) with lower membrane permeability than AMPH, the 4-OH-AMPH metabolite might be more likely than AMPH to reach high levels in the cytoplasm to serve as an effective MAO inhibitor.
