The present study shows that 5-HT exerts a direct influence on Aspergillus spp., as ergosterol synthesis, fungal membrane integrity, germination and hyphal elongation (P < 0.05) were significantly decreased. Moreover, 5-HT was found to accumulate intracellularly, independent of incubation time, temperature and metabolic energy. Short exposure of 5-HT resulted in a lag of re-growth in several fungi.
Many potential targets have been identified in fungi, and antimycotic drugs can act via extracellular and intracellular pathways [2
]. Most currently available drugs inhibit the synthesis of or interact with ergosterol, or directly damage the fungal cell membrane [22
]. In our study, 5-HT exerted a dose-dependent decrease of ergosterol synthesis (P
< 0.05) followed by loss of cell membrane integrity in Aspergillus
spp. The decrease in fungal viability is in parallel with an increase in intracellular fluorescence due to 5-HT accumulation; 5-HT internalisation may therefore result from cell membrane leakage in fungi.
The plasma membrane is a dynamic structure that segregates the intracellular milieu from the extracellular environment by regulating the entry and exit of small and large molecules [23
]. It has been shown that active transport of macromolecules, peptides or proteins into hyphae of various other filamentous fungi can be stopped by inhibitors of oxidative phosphorylation [14
]. Our data show that 5-HT internalisation was not prevented in the presence of inhibitors of oxidative phosphorylation. Neither KCN nor temperature treatment prevented the entry of 5-HT into the hyphal cytoplasm. These data refute active 5-HT transport into the fungal cytoplasm.
The ability of Aspergillus
spp. to undergo morphological changes is an important virulence factor [24
], as the onset of infection is associated with the appearance of hyphae. 5-HT altered the morphology of Aspergillus
spp., since hyphal elongation was significantly decreased and a lack of germination was seen in some isolates. Moreover, a lag of re-growth was observed after exposure to 5-HT and the extent of these effects was dependent on the concentration and incubation time. The maximum duration of lag of re-growth was observed at incubation times shorter than those required for previous studies [25
]. A similar result was found for sertraline, a selective serotonin reuptake inhibitor [27
Currently, the role of 5-HT in antifungal host defence is unclear. There is a coincidence of low 5-HT in certain diseases, e.g. AIDS and an increased rate of infections [12
]. In vivo, the brain and enterochromaffin cells are the main producers of 5-HT, with 5-HT release from enterochromaffin cells taken up by and stored predominantly in platelets and mast cells [6
]. 5-HT is stored in dense granules of platelets at 65mM and is released by platelet degranulation [28
]. We identified fungal killing at concentrations between 14.6mM and 58mM [10
] as well as alterations in candidal virulence at 3μM [25
]. Base levels of 5-HT in serum are ca. 3.5μM [29
] and arise in multiple pathological situations. Our data indicate that 5-HT had only a weak influence on aspergilli at low concentrations but strong effects at pathological levels. The role of 5-HT release from platelets in defence against Aspergillus
spp. needs to be further investigated, as we and others observed that platelets can damage hyphae of Aspergillus
In conclusion, our data indicate that 5-HT accumulates intracellularly, independent of time, temperature and energy. The interaction of 5-HT with Aspergillus spp. affects hyphal growth and diminishes fungal cell membrane integrity in vitro.