| Summary: | Since sildenafil's (Viagra®), a phospodiesterase type 5 (PDE5) inhibitor, approval for the
treatment of male erectile dysfunction (MED) in the United States early 1998, 274
adverse event reports were filed by the Food and Drug Administration (FDA) between 4
Jan. 1998 and 21 Feb. 2001 with sildenafil as the primary suspect of various
neurological disturbances, including amnesia and aggressive behaviour (Milman and
Arnold, 2002). These and other research findings have prompted investigations into the
possible central effects of sildenafil.
The G protein-coupled muscarinic adetylcholine receptors (mAChRs) and serotonergic
receptors (5HT-Rs), have been linked to antidepressant action (Brink et al. 2004).
GPCRs signal through the phosphatidylinositol signal transduction pathway known to
activate protein kinases (PKs). Since the nitric oxide (NO)-guanylyl cyclase signal
transduction pathway is also known to involve the activation of PKs (via cyclic guanosine
monophosphate (cGMP)), the scope is opened for sildenafil to possibly modulate the
action of antidepressants by elevating cGMP levels.
It is generally assumed that excitotoxic delayed cell death is pathologically linked to an
increase in the release of excitatory neurotransmitters e.g. glutamate. Glutamate
antagonists, especially those that block the define NMDA-receptors, are neuroprotective,
showing the importance of the NMDA-NO-cGMP pathway in neuroprotection (Brandt et
al., 2003). Sildenafil may play a role in neuroprotection by elevating cGMP levels.
Aims: The aims of the study were to investigate any neuroprotective properties of
sildenafil, as well as modulating effects of sildenafil pre-treatment on mAChR function.
Methods: Human neuroblastoma SH-SY5Y or human epithelial HeLa cells were seeded
in 24-well plates and pre-treated for 24 hours in serum-free medium with no drug
(control), PDE5 inhibitors sildenafil (100nM and 450 nM), dipiridamole (20 µM) or
zaprinast (20 µM), non-selective PDE inhibitor 3-isobutyl-I-methylxanthine (IBMX -
ImM), cGMP analogue N2,2'-0-dibutyrylguanosine 3'5'-cyclic monophosphate sodium
salt (500 µM), guanylcyclase inhibitor 1H-[1 ,2,4]oxadiazolo[4,3-a]quinoxalin-I-one (ODQ
- 3 µM) or sildenafil + ODQ (450 nM and 3 µM respectively). Thereafter cells were used
to determine mAChR function by constructing dose-response curves of methacholine or
to determine cell viability utilising the Trypan blue, propidium iodide and MTT tests for
cell viability.
Results: Sildenafil pre-treatments induced a 2.5-fold increase in ,the Emax value of
methacholine in neuronal cells but did not show a significant increase in epithelial cells
The Trypan blue test suggests that neither the PDE5 inhibitors nor a cGMP analogue
show any neuroprotection. Rather, sildenafil 450 nM, dipiridamole and IBMX displayed
a neurodegenerative effect. The MTT test was not suitable, since pre-treatment with the
abovementioned drugs inhibited the formation of forrnazan. The propidium iodide assay
could also not be used, due to severe cell loss.
Conclusion: Sildenafil upregulates mAChR function in SH-SY5Y cells and displays a
neurodegenerative, and not a protective property, in neuronal cells. This is not likely to
be associated with its PDE5 inhibitory action, but may possibly be linked to an increase
in cGMP levels via the NO-cGMP pathway. === Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2005.
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