Effects of TSPO-ligands on mitochondrial volume and mitochondrial by htt39969


									                                     Effects of TSPO-ligands
                  on mitochondrial volume and mitochondrial membrane potential
                  Lara Testai 1, Maria Cristina Breschi1, Federico Da Settimo2, Alma Martelli1,
                          Francesca Simorini2, Sabrina Taliani2, Vincenzo Calderone1
  Dipartimento di Psichiatria, Neurobiologia, Farmacologia e Biotecnologie, Università di Pisa, via Bonanno, 6,
             Dipartimento di Scienze Farmaceutiche, Università di Pisa, via Bonanno, 6, 56120-PISA

Background- Mitochondria are deeply involved in the processes underlying cellular survival/death. They play a
decisive role to establish the cell destiny, controlling either apoptotic or necrotic pathways through the
modulation of a mitochondrial multi-protein complex, known as mitochondrial permeability transition pore
(MPTP). The extensive and prolonged opening of this pore causes the release of pro-apoptotic inter-membrane
proteins and the dissipation of mitochondrial membrane potential (Δψ). The translocator protein (TSPO),
previously classified as peripheral benzodiazepine receptor, is a constituent of MPTP in association with other
mitochondrial proteins. Although the dynamics undergoing the MPTP modulation are still largely unknown, it is
recognised that TSPO ligands may act as potential therapeutic agents useful in the treatment of a number of
pathological conditions, including cancer, ischemia-reperfusion injury, inflammation processes and immune
responses [1]. In these perspectives, a new class of indolylglyoxylamide TSPO ligands has been recently
developed [2-3].
Aim- In order to evaluate the effects of this new chemo-type of TSPO ligands on mitochondrial functions, six
indolylglyoxylamide compounds (1-6) have been selected and tested on rat hepatic mitochondria. In binding
assays on rat kidney membranes by competition experiments against 3[H]-PK11195, four of them (1, 2, 4 and 6)
exhibited high affinity vs TSPO, while compounds 3 and 5 were almost ineffective.
                                                           Compds         R1             R2           R3     R4
                         N                                   1         (CH2)3CH3      (CH2)3CH3       F      H
              O                                              2         (CH2)5CH3      (CH2)5CH3       Cl     H
                                                             3         (CH2)2CH3      (CH2)2CH3       H     OCH3
                                                             4         (CH2)3CH3      (CH2)3CH3       Cl     Cl
                                                             5           CH3           CH2CH3         H      H
                                                             6          CH2CH3        (CH2)3CH3       Cl     Cl

Results and Discussion- Compounds 1, 2, 4, 6 (30μM) per se did not induce any significant change of the
mitochondrial volume (recorded spectrophotometrically at 520 nm), but markedly reduced the mitochondrial
swelling induced by addition of CaCl2 200 µM. Compounds 3 and 5, devoid of TSPO affinity, did not influence
the Ca++-induced swelling.
Moreover, the administration of compounds 1, 2, 4, 6, but not of 3 and 5, at the mitochondrial suspension
induced an evident and concentration-dependent membrane depolarization, potentiometrically measured by
TPP+-sensitive micro-electrodes.
Because of the close correlation between the data emerging from these experimental procedures and those
previously reported in the binding assays [2-3], it can be hypothesised that the interaction with TSPO is the first
step of the chain of molecular mechanisms accounting for the functional effects on mitochondrial matrix volume
and on Δψ. Further studies will be focused to understand the downstream electrophysiological and/or metabolic
steps responsible for these functional effects.

[1] Veenman L, Gavish M, Pharmacol. Ther. 2006, 10: 503-24
[2] Primofiore G, Da Settimo F, Taliani S, Simorini F, Patrizi MP, Novellino E, et al., J. Med. Chem. 2004, 47:
[3] Da Settimo F, Simorini F, Taliani S, La Motta C, Marini AM, Salerno S, et al., J. Med. Chem. 2008, 51:

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