Antidiabetic Effect of a New Original NT-3 Dipeptide Mimetic.

It was previously established that the original dipeptide mimetic of the 4th loop of NT-3, hexamethylenediamide bis-(N-monosuccinyl-L-asparaginyl-L-asparagine) (GTS-301), has a pronounced neuroprotective effect in vitro at concentrations of 10-5-10-12 М. In the present study, experiments on the streptozotocin-induced diabetes model in C57Bl/6 mice showed that GTS-301, when administered intraperitoneally for 32 days at doses of 0.1 and 0.5 mg/kg, has antidiabetic activity manifested in a reduction of hyperglycemia and polydipsia and in an increase in animal survival. The results obtained confirm the concept of the similarity of neurochemical mechanisms underlying the regulation of functions of neurons and ß-cells.

Low-Molecular-Weight Perorally Active Nerve Growth Factor Mimetic Reduces Manifestations of Diabetic Neuropathy in Wistar Rats.

A low-molecular-weight nerve growth factor mimetic, compound GK-2 (bis-(N-monosuccinyl- L-glutamyl-L-lysine)hexamethylenediamide) that previously demonstrated antidiabetic activity in rats with streptozotocin-induced type 2 diabetes mellitus was studied on the model of diabetic neuropathy. It was found that in 8 weeks after diabetes mellitus development, untreated diabetic rats demonstrated impaired tactile sensitivity in von Frey test, while GK-2 therapy (7.5 mg/kg orally for 28 days) restored this parameter. The decrease of tactile sensitivity in diabetic neuropathy closely correlated with the severity of hyperglycemia (r=0.76). Our findings are consistent with the concept on the role of glucose toxicity and nerve growth factor deficiency in the pathogenesis of diabetic neuropathy and attest to feasibility of further studies of nerve growth factor mimetic GK-2 as a potential treatment for diabetes and diabetic neuropathy.

Analysis of the Role of PI3K/Akt Pathway in the Realization of the Normalizing Effect of Low-Molecular-Weight Mimetics of NGF and BDNF on Activity of Prooxidant and Antioxidant Systems in C57BL/6 Mice with Modeled Diabetes.

The effect of low-molecular-weight mimetics of NGF and BDNF (GK-2 and GSB-214 in a dose 0.5 mg/kg, respectively) on malondialdehyde content and activity of an antioxidant defense enzyme glutathione peroxidase was studied in experiments on C57BL/6 mice with streptozotocin-induced diabetes. An increase in the malondialdehyde content indicating enhanced formation of peroxidation products and a decrease of glutathione peroxidase activity in the blood plasma of untreated diabetic animals were revealed. Both studied mimetics were shown to attenuate the severity of these disorders. Since the ability of these compounds to activate the PI3K/Akt signaling pathway was previously demonstrated in vitro on HT-22 cell culture, we studied the effect of LY294002, an inhibitor of this pathway, on the above parameters. It was found that LY294002 attenuates the normalizing effect of GK-2 and GSB-214 only in relation to glutathione peroxidase activity, but not malondialdehyde level.

The PI3K/Akt Cascade Is Involved in the Antidiabetic Effect of Compound GSB-214, a Low-Molecular-Weight BDNF Mimetic.

In our previous studies on the streptozotocin model of diabetes we hypothesized that activation of the PI3K/Akt signaling pathway is essential for the realization of the antidiabetic effect of low-molecular-weight NGF and BDNF mimetics. Here we analyze the effect of a specific PI3K/Akt pathway inhibitor (LY 294002) on the antidiabetic effect of the BDNF loop 1 mimetic GSB-214. The experiments on C57BL/6 mice with streptozotocin-induced diabetes showed that GSB-214 attenuated the hyperglycemic effect of streptozotocin and prevented weight loss typical of diabetes, while LY 294002 eliminated these effects of GSB-214. These findings clearly demonstrate the involvement of PI3K/Akt pathway in the implementation of the effects of this low-molecular-weight BDNF mimetic.

Analysis of Cytoprotective Properties of Afobazole in Streptozotocin Model of Diabetes.

Previous in vitro and in vivo studies revealed the neuroprotective effect of anxiolytic Afobazole. Based on similarities in the regulation of functions of neurons and ß cells, we studied the effect of Afobazole on streptozotocin (STZ) model of type 2 diabetes in Wistar rats. Immunohistochemical analysis showed that the decrease in the number of ß cells and a violation of their morphological structure caused by STZ were significantly alleviated by Afobazole administration (10 mg/kg orally for 28 days) to diabetic animals. A correlation between morphometric data and blood glucose level was revealed. A possible role of σ1-receptors in the cytoprotective effects of Afobazole in respect to pancreatic ß cells is discussed.

Drug with Neuroprotective Properties Noopept Does Not Stimulate Cell Proliferation.

Effects of Noopept (N-phenylacetyl-L-prolylglycine ethyl ester) on the relative level of proliferation marker Ki-67 and cell cycle parameters were studied in HEK293 and SH-SY5Y cell lines. The previously established multifactorial mechanism of action of the drug includes enhancement of neurotrophin NGF and BDNF expression and increase in HIF-1 activity. The possible mitogenic action of Noopept was estimated by its effect on cell proliferation. Noopept did not affect cell distribution over G1, S, G2 cell cycle phases and the relative level of proliferation marker Ki-67 in the cell lines under study. These data suggest that Noopept does not stimulate cell growth.

Neuroprotective Dipeptide Noopept Prevents DNA Damage in Mice with Modeled Prediabetes.

In experiments on BALB/c mice, prediabetes was modeled by administration of streptozotocin in a dose of 130 mg/kg. DNA damage was assessed by the method of DNA comets. Noopept (0.5 mg/kg intraperitoneally) was administered for 14 days before and for 6, 13, or 14 days after streptozotocin administration. Despite moderate hyperglycemia and increased malondialdehyde level, the intensity of DNA damage in cells of the pancreas, liver, and kidneys significantly surpassed the control values. Noopept normalized these parameters due to its pronounced antigenotoxic effect. For both the damaging effect of streptozotocin and the normalizing effect of Noopept, DNA changes manifested mainly in terms of atypical DNA comets. Our findings confirm the role of DNA damage in the pathogenesis of diabetes. They indicate the possibility of pharmacological protection of pancreatic ß cells with the neuroprotective drug and provide an important argument in favor of the hypothesis about the similarity of the mechanisms of formation of the resistance of neurons and ß cells to the cytotoxic influences.

Neuroprotective Lithium Salts Protect Pancreatic ß-Сells from Damage.

We studied antidiabetic effects and cytoprotective activity of two lithium salts (lithium chloride and lithium carbonate) on the model of streptozotocin-induced diabetes mellitus type 2 in Wistar rats. Using the method of ß-cells detection with antibodies to insulin, we demonstrated that streptozotocin reduced the number of ß-cells and impaired their morphological structure. Both lithium preparations administered to diabetic animals for 28 days in doses of 10 and 8.9 mg/kg, respectively, attenuated the damaging effect of streptozotocin. This cytoprotective effect of lithium salts manifested in weakening of hyperglycemia, polyphagia, polydipsia, and weight loss. A satisfactory correlation between the morphometric data and blood glucose levels was revealed. The mechanisms of the multitarget action of lithium salts are discussed.

Antidiabetic Properties of Low-Molecular-Weight BDNF Mimetics Depend on the Type of Activation of Post-Receptor Signaling Pathways.

Reduced proliferation and enhanced apoptosis of ß cells in diabetes mellitus are associated with a deficiency of brain-derived neurotrophic factor (BDNF). Low-molecular weight compounds similar to different BDNF loops were synthesized at the V. V. Zakusov Research Institute of Pharmacology. They produce a potentiating effect on TrkB phosphorylation, but differently activate post-receptor signaling pathways. We compared their effects on the severity of streptozotocin-induced diabetes mellitus in C57Bl/6 mice. The antidiabetic effect (estimated from the degree of hyperglycemia and dynamics of body weight) was typical of GSB-214 compound that selectively activates PI3K/Akt. This activity was not revealed in GTS-201, selective activator of MAPK/Erk. GSB-106 compound activating both signaling pathways exhibited weak antidiabetic activity. Our results indicate that the antidiabetic effect is mainly related to activation of the PI3K/Akt signaling pathway.

Antidiabetic Activity of Afobazole in Wistar Rats.

Using the streptozotocin model of type 2 diabetes mellitus in Wistar rats, we compared antidiabetic activity of anxiolytic Afobazole with that of metformin. Afobazole in a dose of 10 mg/kg reduced streptozotocin-induced hyperglycemia and polyphagia and prevented accumulation of malonic dialdehyde, being not inferior to metformin in a dose of 300 mg/kg, and was even more effective than metformin in body weight recovery, elimination of polydipsia, and preservation of these effects after treatment withdrawal.

Nootropic dipeptide noopept enhances inhibitory synaptic transmission in the hippocampus.

Application of nootropic agent Noopept on hippocampal slices from Wistar rats enhanced the inhibitory component of total current induced by stimulation of Shaffer collaterals in CA1 pyramidal neurons, but did not affect the excitatory component. A direct correlation between the increase in the amplitude of inhibitory current and agent concentration was found. The substance did not affect the release of inhibitory transmitters from terminals in the pyramidal neurons, which indicated changes in GABAergic interneurons.

Comparative activity of proline-containing dipeptide noopept and inhibitor of dipeptidyl peptidase-4 sitagliptin in a rat model of developing diabetes.

Developing diabetes was modeled on adult male Wistar rats by repeated intraperitoneal injections of streptozotocin in a subdiabetogenic dose of 30 mg/kg for 3 days. Proline-containing dipeptide drug Noopept or a standard diabetic drug dipeptidyl peptidase-4 inhibitor sitagliptin was administered per os in a dose of 5 mg/kg before each injection of the toxin and then for 16 days after streptozotocin course. In active control group, spontaneously increase glucose level and reduced tolerance to glucose load (1000 mg/kg intraperitoneally) were observed on the next day after the third administration of toxin. Basal glucose level decreased by day 16, but glucose tolerance remained impaired. Noopept normalized the basal blood glucose level and tolerance to glucose load on the next day after administration of streptozotocin. The effect of Noopept persisted to the end of the experiment. At early terms of the experiment, sitagliptin was somewhat superior to Noopept by the effect on baseline glucose level, but was inferior by the influence on glucose tolerance.. By the end of the experiment, Noopept significantly (by 2 times) surpassed sitagliptin by its effect on glucose tolerance.

Noopept normalizes parameters of the incretin system in rats with experimental diabetes.

Experiments on adult Wistar rats with streptozotocin-induced diabetes showed that antihyperglycemic activity of an original nootropic and neuroprotective drug Noopept (N-phenylacetyl-L-prolylglycine ethyl ester) is more pronounced under conditions of oral application than after intraperitoneal injection. These data provided a basis for studying the effect of Noopept on major indexes of the incretin system. Streptozotocin was shown to decrease the concentrations of incretin GLP-1 and insulin in the blood. Noopept had a normalizing effect on these parameters. This influence of Noopept was not related to the inhibition of a major enzyme metabolizing incretins (dipeptidyl peptidase IV). A reference drug sitagliptin also increased the contents of incretins and insulin, which was associated with the inhibition of dipeptidyl peptidase IV. It is known that GLP-1 increases NGF expression in the insular system. Our results suggest that the increase in incretin activity contributes to the antiapoptotic effect of Noopept on pancreatic ß cells. The mechanism for an increase in blood GLP-1 level after oral application of Noopept requires further investigations.

Efficiency of noopept in streptozotocin-induced diabetes in rats.

We studied the effects of new nootropic and neuroprotective drug Noopept (N-phenylacetyl-L-prolylglycine ethyl ester) in various dosage regimens on the dynamics of glycemia, body weight, and pain sensitivity in rats receiving diabetogenic toxin streptozotocin. In experimental diabetic rats, Noopept alleviated glycemia and weight loss and normalized enhanced pain sensitivity. The normalizing effect of Noopept was most pronounced when it was administered as a preventive agent prior to injection of the toxin. Both preventive and therapeutic administration of Noopept (delayed injections included) significantly weakened the examined metabolic effects of diabetogenic toxin. Possible mechanisms of the antidiabetic action of Noopept are analyzed.

Effects of neurotensin dipeptide analog dilept on dopamine metabolism and synthesis in the nucleus accumbens of Wistar rats.

We studied the effects of neurotensin dipeptide analog Dilept (N-caproyl-L-prolyl-L-tyrosine methyl ester) on dopamine metabolism and synthesis in the nucleus accumbens of Wistar rats. Dilept increased the levels of dopamine and its metabolites (homovanillic acid and dioxyphenylalanine) and stimulated dopamine turnover in this structure. Dilept accelerated dopamine synthesis under conditions of pulsed activity blockade in dopaminergic neuron by injection of γ-butyrolactone combined with inhibition of aromatic acid decarboxylase with 3-hydroxybenzylhydrazine. The spectrum of pharmacological activities of Dilept towards the dopaminergic system of the nucleus accumbens was similar to that of atypical neuroleptics and neurotensin (endogenous antipsychotic).

Genotype-dependent characteristics of behavior in mice in cognitive tests. The effects of Noopept.

Male C57BL/6J, BALB/c, and DBA/2J mice showed differences in their abilities to perform two cognitive tests. C57BL/6J mice had good learning ability and memory trace retention (at 10 days) in a simplified Morris maze, while BALB/c mice had low levels of memory trace retention and DBA/2J mice had low learning ability in this test. I.p. administration of the nootropic agent Noopept (GVS-111, N-phenylacetyl-L-prolylglycine ethyl ester) at a dose of 0.5 mg/kg 15 min before the start of the test induced significant improvements in long-term memory in this test in BALB/c mice but no further improvement in C57BL/6J mice, and had no effect in DBA/2J mice. On testing the ability to extrapolate the direction of movement of a stimulus, administration of Noopept increased the proportion of correct responses in C57BL/6J and BALB/c mice, but had no effect in DBA/2J mice.

A Novel Dipeptide NGF Mimetic GK-2 Selectively Activating the PI3K/AKT Signaling Pathway Promotes the Survival of Pancreatic ß-Cells in a Rat Model of Diabetes.

We investigated the cytoprotective effect of a novel low-molecular-weight NGF mimetic, GK-2 (hexamethylenediamide bis-N-monosuccinyl-L-glutamyl-L-lysine), on pancreatic ß-cells. The neuroprotective effect of GK-2 had been previously shown to be associated with selective activation of the PI3K/Akt signaling pathway. In this study, rats with streptozotocin (STZ)-induced type 2 diabetes mellitus were used. Metformin was used as a reference drug. STZ was immunohistochemically demonstrated to reduce the number of ß-cells and affect their morphological structure. Treatment of diabetic animals with GK-2 (at a dose of 0.5 mg/kg intraperitoneally or 5 mg/kg orally) or metformin (300 mg/kg orally) for 28 days reduced the damaging effect of STZ. The effect of GK-2 on manifestations of STZ-induced diabetes, such as hyperglycemia, weight loss, polyphagia, and polydipsia, was comparable to that of metformin, while the cytoprotective activity of GK-2 was slightly stronger than that of metformin. A strong positive correlation between morphometric parameters and the blood glucose level was revealed. The GK-2 cytoprotective effect on ß-cells is supposed to manifest through the PI3K/Akt signaling pathway.

Low-Molecular-Weight NGF Mimetic Corrects the Cognitive Deficit and Depression-like Behavior in Experimental Diabetes.

Based on the comorbidity of diabetes, depression, and dementia and recognizing that a deficiency of the nerve growth factor (NGF) is involved in all of these kinds of pathologies, we studied the effect of the mimetic of dimeric dipeptide NGF loop 4, GK-2, on a model of streptozotocin-induced type 2 diabetes in C57Bl/6 mice. GK-2 [hexamethylenediamide bis-(N-monosuccinyl-glutamyl-lysine)] was synthesized at the V.V. Zakusov Scientific Research Institute of Pharmacology. The study revealed the ability of GK-2 to ameliorate hyperglycemia induced by streptozotocine (STZ 100 mg/kg i.p.) in C57Bl/6 mice, to restore learning ability in the Morris Water Maze test, and to overcome depression after both intraperitoneal (0.5 mg/kg) and peroral (5 mg/kg) long-term administration. The presence of the listed properties and their preservation in the case of peroral treatment determines the prospects of research. Taking into account the previous findings on the ability of GK-2 to selectively activate PI3K/Akt, these data suggest that Akt-signaling is sufficient for pancreatic beta cell function. GK-2 has been shown to exhibit pronounced neuroprotective activity. The coexistence of neuroprotective and antidiabetic effects is in agreement with the fundamental concept holding that the function of neurons and pancreatic beta cells is controlled by similar mechanisms.

Molecular Mechanism Underlying the Action of Substituted Pro-Gly Dipeptide Noopept.

This study was performed in order to reveal the effect of Noopept (ethyl ester of N-phenylacetyl-Lprolylglycine, GVS-111) on the DNA-binding activity of transcriptional factors (TF) in HEK293 cells transiently transfected with luciferase reporter constructs containing sequences for CREB, NFAT, NF-κB, p53, STAT1, GAS, VDR, HSF1, and HIF-1. Noopept (10 µM) was shown to increase the DNA-binding activity of HIF-1 only, while lacking the ability to affect that of CREB, NFAT, NF-κB, p53, STAT1, GAS, VDR, and HSF1. Noopept provoked an additional increase in the DNA-binding activity of HIF-1 when applied in conditions of CoCl2-induced HIF- 1 stabilization. The degree of this HIF-positive effect of Noopept was shown to be concentration-dependent. Piracetam (1 mM) failed to affect significantly any of the TF under study. The results of molecular docking showed that Noopept (L-isomer), as well as its metabolite, L-isomer of N-phenyl-acetylprolyl, unlike its pharmacologically ineffective D-isomer, is able to bind to the active site of prolyl hydroxylase 2. Taking into account the important role of the genes activated by HIF-1 in the formation of an adaptive response to hypoxia, data on the ability of Noopept to provoke a selective increase in the DNA-binding activity of HIF-1 explain the wide spectrum of neurochemical and pharmacological effects of Noopept revealed before. The obtained data allow one to propose the HIF-positive effect as the primary mechanism of the activity of this Pro-Gly-containing dipeptide.

Identification of a novel endogenous memory facilitating cyclic dipeptide cyclo-prolylglycine in rat brain.

Using high-performance liquid chromatography, gas-chromatography and chromato-mass spectrometry methods a novel endogenous cyclic dipeptide cyclo-prolylglycine was identified in rat brain. Its content according to gas chromatography is 2.8 +/- 0.3 nmol/g wet brain. Synthetic cyclo-prolylglycine has demonstrated antiamnesic activity in the passive avoidance test in rats at a dose of 0.1 mg/kg i.p. Cyclic dipeptide cyclo-prolylglycine seems to be a memory facilitating substance and its presence in rat brain suggests the existence of a new mechanism of memory regulation.