Ischemic tolerance determines resistance to lethal ischemia gained by a prior sublethal stimulus (i.e., preconditioning). We reproduced this effect in two variants. In vitro the preliminary short (5 s) photodynamic treatment (PDT) (photosensitizer Photosens, 10 nM, 30 min preincubation; laser: 670 nm, 100 mW/cm2) significantly reduced the necrosis of neurons and glial cells in the isolated crayfish stretch receptor, which was caused by following 30-min PDT by 66% and 46%, respectively. In vivo PDT of the rat cerebral cortex with hydrophilic photosensitizer Rose Bengal (i.v. administration, laser irradiation: 532 nm, 60 mW/cm2, 3 mm beam diameter, 30 min) caused occlusion of small brain vessels and local photothrombotic infarct (PTI). It is a model of ischemic stroke. Cerebral tissue edema and global necrosis of neurons and glial cells occurred in the infarction core, which was surrounded by a 1.5 mm transition zone, penumbra. The maximal pericellular edema, hypo- and hyperchromia of neurons were observed in penumbra 24 h after PTI. The repeated laser irradiation of the contralateral cerebral cortex also caused PTI but lesser as compared with single PDT. Preliminary unilateral PTI provided ischemic tolerance: at 14 day after second exposure the PTI volume significantly decreased by 24% than in the case of a single exposure. Sensorimotor deficits in PDT-treated rats was registered using the behavioral tests. The preliminary PTI caused the preconditioning effect.
Photodynamic therapy (PDT) is currently used in the treatment of brain tumors. However, not only malignant cells but also neighboring normal neurons and glial cells are damaged during PDT. In order to study the potential role of transcription factors—nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), activator protein (AP-1), and signal transducer and activator of transcription-3 (STAT-3)—in photodynamic injury of normal neurons and glia, we photosensitized the isolated crayfish mechanoreceptor consisting of a single sensory neuron enveloped by glial cells. Application of different inhibitors and activators showed that transcription factors NF-κB (inhibitors caffeic acid phenethyl ester and parthenolide, activator betulinic acid), AP-1 (inhibitor SR11302), and STAT-3 (inhibitors stattic and cucurbitacine) influenced PDT-induced death and survival of neurons and glial cells in different ways. These experiments indicated involvement of NF-κB in PDT-induced necrosis of neurons and apoptosis of glial cells. However, in glial cells, it played the antinecrotic role. AP-1 was not involved in PDT-induced necrosis of neurons and glia, but mediated glial apoptosis. STAT-3 was involved in PDT-induced apoptosis of glial cells and necrosis of neurons and glia. Therefore, signaling pathways that regulate cell death and survival in neurons and glial cells are different. Using various inhibitors or activators of transcription factors, one can differently influence the sensitivity and resistance of neurons and glial cells to PDT.
Photodynamic therapy (PDT) effect is a promising adjuvant modality for diagnosis and treatment of brain cancer. It is of importance that the bright fluorescence of most photosensitizers provides visualization of brain tumors. This is successfully used for fluorescence-guided tumor resection according to the principle “to see and to treat.” Non-oncologic application of PDT effect for induction of photothrombotic infarct of the brain tissue is a well-controlled and reproducible stroke model, in which a local brain lesion is produced in the predetermined brain area. Since normal neurons and glial cells may also be damaged by PDT and this can lead to unwanted neurological consequences, PDT effects on normal neurons and glial cells should be comprehensively studied. We overviewed the current literature data on the PDT effect on a range of signaling and epigenetic proteins that control various cell functions, survival, necrosis, and apoptosis. We hypothesize that using cell-specific inhibitors or activators of some signaling proteins, one can selectively protect normal neurons and glia, and simultaneously exacerbate photodynamic damage of malignant gliomas.
Radachlorin is a chlorine-derivative photosensitizer used currently in photodynamic therapy (PDT). We studied photodynamic effect of Radachlorin on neurons and surrounding glial cells. The crayfish abdominal stretch receptor and abdominal nerve cord of were used as models. Radachlorin absorption spectrum contained maximums at 420 and 654 nm and fluorescence maximum at 664 nm. In the crayfish stretch receptor Radachlorin localized predominantly to the glial envelope and penetrated only slightly into the neuron. Radachlorin accumulated rapidly in the nerve cord tissue within 30 min. Its elimination in the dye-free solution occurred slower: 11% loss during 2h. Radachlorin-PDT inactivated the neuron, induced necrosis of neurons and glial cells and glial apoptosis at concentrations as low as 10-10-10-9 M. rapidly accumulates in the nervous tissue, mainly in glial cells. High photodynamic efficacy, the adsorption spectrum with the red maximum within the “transparency window," rapid accumulation and elimination characterize Radachlorin as a promising photosensitizer for photodynamic therapy.
Photodynamic therapy (PDT) is used for selective destruction of cells, in particular, for treatment of brain tumors. However, photodynamic treatment damages not only tumor cells, but also healthy neurons and glial cells. To study the possible role of NF-κB in photodynamic injury of neurons and glial cells, we investigated the combined effect of photodynamic treatment and NF-κB modulators: activator betulinic acid, or inhibitors parthenolide and CAPE on an isolated crayfish stretch receptor consisting of a single neuron surrounded by glial cells. A laser diode (670 nm, 0.4 W/cm2) was used as a light source. The inhibition of NF-κB during PDT increased the duration of neuron firing and glial necrosis and decreased neuron necrosis and glial apoptosis. The activation of NF-κB during PDT increased neuron necrosis and glial apoptosis and decreased glial necrosis. The difference between the effects of NF-κB modulators on photosensitized neurons and glial cells indicates the difference in NF-κB-mediated signaling pathways in these cell types. Thus, NF-κB is involved in PDT-induced shortening of neuron firing, neuronal and glial necrosis, and apoptosis of glial cells.
Nitric oxide (NO) is an important second messenger, involved in the implementation of various cell functions. It regulates various physiological and pathological processes such as neurotransmission, cell responses to stress, and neurodegeneration. NO synthase is a family of enzymes that synthesize NO from L-arginine. The activity of different NOS isoforms depends both on endogenous and exogenous factors. In particular, it is modulated by oxidative stress, induced by photodynamic therapy (PDT). We have studied the possible role of NOS in the regulation of survival and death of neurons and surrounding glial cells under photo-oxidative stress induced by photodynamic treatment (PDT). The crayfish stretch receptor consisting of a single identified sensory neuron enveloped by glial cells is a simple but informative model object. It was photosensitized with alumophthalocyanine photosens (10 nM) and irradiated with a laser diode (670 nm, 0.4 W/cm2). Antinecrotic and proapoptotic effects of NO on the glial cells were found using inhibitory analysis. We have shown the role of inducible NO synthase in photoinduced apoptosis and involvement of neuronal NO synthase in photoinduced necrosis of glial cells in the isolated crayfish stretch receptor. The activation of NO synthase was evaluated using NADPH-diaphorase histochemistry, a marker of neurons expressing the enzyme. The activation of NO synthase in the isolated crayfish stretch receptor was evaluated as a function of time after PDT. Photodynamic treatment induced transient increase in NO synthase activity and then slowly inhibited this enzyme.
Photodynamic treatment that causes intense oxidative stress and kills cells is currently used in neurooncology. However,
along with tumor it damages surrounding healthy neurons and glial cells. In order to study the possible role of
glutamate-related signaling pathways in photodynamic injury of neurons and glia, we investigated photodynamic effect
of alumophthalocyanine Photosens on isolated crayfish stretch receptor that consists of a single neuron surrounded by
glial cells. The laser diode (670 nm, 0.4 W/cm2) was used for dye photoexcitation. Application of glutamate increased
photodynamically induced necrosis of neurons and glial cells but significantly decreased glial apoptosis. The natural
neuroglial mediator N-acetylaspartylglutamate, which releases glutamate after cleavage in the extracellular space by
glutamate carboxypeptidase II, also inhibited photoinduced apoptosis. Inhibition of glutamate carboxypeptidase II,
oppositely, enhanced apoptosis of glial cells. These data confirm the anti-apoptotic activity of glutamate. Application of
NMDA or inhibition of NMDA receptors by MK801 did not influence photodynamic death of neurons and glial cells
that indicated nonparticipation of NMDA receptors in these processes. Inhibition of metabotropic glutamate receptors
by AP-3 decreased PDT-induced apoptosis. One can suggest that crayfish neurons naturally secrete NAAG, which being
cleaved by GCOP produces glutamate. Glutamate prevents photoinduced apoptosis of glial cells possibly through
metabotropic but not ionotropic glutamate receptors.
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