[+]-Huperzine A treatment protects against N-methyl-d-aspartate-induced seizure/status epilepticus in rats

The toxicity of organophosphorous (OP) nerve agents is attributed to their irreversible inhibition of acetylcholinesterase (AChE), which leads to excessive accumulation of acetylcholine (ACh) and is followed by the release of excitatory amino acids (EAA). EAAs sustain seizure activity and induce neuropathology due to over-stimulation of N-methyl-d-aspartate (NMDA) receptors. Huperzine A (Hup A), a blood–brain barrier permeable selective reversible inhibitor of AChE, has been shown to reduce EAA-induced cell death by interfering with glutamate receptor-gated ion channels in primary neuronal cultures. Although [−]-Hup A, the natural isomer, inhibits AChE approximately 38-fold more potently than [+]-Hup A, both [−]- and [+]-Hup A block the NMDA channel similarly. Here, we evaluated the protective efficacy of [+]-Hup A for NMDA-induced seizure in a rat model. Rats implanted with radiotelemetry probes to record electroencephalography (EEG), electrocardiography (ECG), body temperature, and physical activity were administered various doses of [+]-Hup A (intramuscularly) and treated with 20 μg/kg NMDA (intracerebroventricular) 20–30 min later. For post-exposure, rats were treated with [+]-Hup A (3 mg/kg, intramuscularly) 1 min after NMDA (20 μg/kg). Our data showed that pre- and post-exposure, [+]-Hup A (3 mg/kg) protects animals against NMDA-induced seizures. Also, NMDA-administered animals showed increased survival following [+]-Hup A treatment. [+]-Hup A has no visible effect on EEG, heart-rate, body temperature, or physical activity, indicating a reduced risk of side effects, toxicity, or associated pathology. Our results suggest that [+]-Hup A protects against seizure and status epilepticus (SE) by blocking NMDA-induced excitotoxicity in vivo. We propose that [+]-Hup A, or a unique combination of [+]- and [−]-Hup A, may prove to be effective for pre- and post-exposure treatment of lethal doses of OP-induced neurotoxicity.     

+]-Huperzine A treatment protects against N-methyl-D-aspartate-induced seizure/status epilepticus in rats

The toxicity of organophosphorous (OP) nerve agents is attributed to their irreversible inhibition of acetylcholinesterase (AChE), which leads to excessive accumulation of acetylcholine (ACh) and is followed by the release of excitatory amino acids (EAA). EAAs sustain seizure activity and induce neuropathology due to over-stimulation of N-methyl-d-aspartate (NMDA) receptors. Huperzine A (Hup A), a blood-brain barrier permeable selective reversible inhibitor of AChE, has been shown to reduce EAA-induced cell death by interfering with glutamate receptor-gated ion channels in primary neuronal cultures. Although [-]-Hup A, the natural isomer, inhibits AChE approximately 38-fold more potently than [+]-Hup A, both [-]- and [+]-Hup A block the NMDA channel similarly. Here, we evaluated the protective efficacy of [+]-Hup A for NMDA-induced seizure in a rat model. Rats implanted with radiotelemetry probes to record electroencephalography (EEG), electrocardiography (ECG), body temperature, and physical activity were administered various doses of [+]-Hup A (intramuscularly) and treated with 20mug/kg NMDA (intracerebroventricular) 20-30min later. For post-exposure, rats were treated with [+]-Hup A (3mg/kg, intramuscularly) 1min after NMDA (20mug/kg). Our data showed that pre- and post-exposure, [+]-Hup A (3mg/kg) protects animals against NMDA-induced seizures. Also, NMDA-administered animals showed increased survival following [+]-Hup A treatment. [+]-Hup A has no visible effect on EEG, heart-rate, body temperature, or physical activity, indicating a reduced risk of side effects, toxicity, or associated pathology. Our results suggest that [+]-Hup A protects against seizure and status epilepticus (SE) by blocking NMDA-induced excitotoxicity in vivo. We propose that [+]-Hup A, or a unique combination of [+]- and [-]-Hup A, may prove to be effective for pre- and post-exposure treatment of lethal doses of OP-induced neurotoxicity.     

Protection of red blood cell acetylcholinesterase by oral huperzine A against ex vivo soman exposure: Next generation prophylaxis and sequestering of acetylcholinesterase over butyrylcholinesterase

As part of a phase Ib clinical trial to determine the tolerability and safety of the highly specific acetylcholinesterase (AChE) inhibitor huperzine A, twelve (12) healthy elderly individuals received an escalating dose regimen of huperzine A (100, 200, 300, and 400 μg doses, twice daily for a week at each dose), with three (3) individuals as controls receiving a placebo. Using the WRAIR whole blood cholinesterase assay, red blood cell AChE and plasma butyrylcholinesterase (BChE) were measured in unprocessed whole blood samples from the volunteers following each dose, and then for up to 48 h following the final and highest (400 μg) dose to monitor the profile of inhibition and recovery of AChE. Significant inhibition of AChE was observed, ranging from 30–40% after 100 μg to >50% at 400 μg, and peaking 1.5 h after the last dose. Gradual recovery of AChE activity then occurs, but even 48 h after the last dose red blood cell AChE was about 10% below control (pre-dose) values. Huperzine A levels in plasma peaked 1.5 h after the final 400 μg dose (5.47 ± 2.15 ng/mL). Plasma BChE was unaffected by huperzine A treatment (as expected).Aliquots of huperzine A-containing (from three individuals) and placebo blood samples were exposed ex vivo to the irreversible nerve agent soman (GD) for 10 min, followed by removal of unbound huperzine and soman from the blood by passing through a small C₁₈ reverse phase spin column. Eluted blood was diluted in buffer, and aliquots taken at various time intervals for AChE and BChE activity measurement to determine the time taken to achieve full return in activity of the free enzyme (dissociation from the active site of AChE by huperzine A), and thus the proportion of AChE that can be protected from soman exposure. Huperzine A-inhibited red blood cell (RBC) AChE activity was restored almost to the level that was initially inhibited by the drug. The increased doses of huperzine A used were well tolerated by these patients and in this ex vivo study sequestered more red blood cell AChE than has been previously demonstrated for pyridostigmine bromide (PB), indicating the potential improved prophylaxis against organophosphate (OP) poisoning.     

Pharmacodynamic Study of FS-0311: A Novel Highly Potent,Selective Acetylcholinesterase Inhibitor

(1) This study was to evaluate the anti-cholinesterase (ChE), cognition enhancing and neuroprotective effects of FS-0311, a bis-huperzine B derivative. (2) ChE activity was evaluated using a spectrophotometric method. Cognitive deficits in mice were induced by scopolamine or transient brain ischemia and reperfusion. Water maze was used to detect the cognitive performance. PC12 cell injury was induced by β-amyloid 25–35 (Aβ_25–35), oxygen-glucose deprivation (OGD), or staurosporine treatment. (3) FS-0311 was a potent, highly specific inhibitor of acetylcholinesterase (AChE). FS-0311 bound to AChE in a reversible manner, causing linear mixed-type inhibition. FS-0311 had a high oral bioavailability and a long duration of AChE inhibitory action in vivo. FS-0311 was found to antagonize cognitive deficits induced by scopolamine or transient brain ischemia and reperfusion in a water maze task. FS-0311 possessed the ability to protect PC12 cells against Aβ_25–35 peptide toxicity, OGD insult and staurosporine-induced apoptosis. The neuroprotective effects of FS-0311 appeared to reflect an attenuation of oxidative stress. (4) With the profile of anti-ChE and neuroprotective activities, FS-0311 might be a promising candidate in neurodegenerative diseases, such as Alzheimer’s disease and Vascular dementia.     

[AuCl4]− and Fe3+/[Fe(CN)6]3− ions-derivated immunosensing interface for electrochemical immunoassay of carcinoembryonic antigen in human serum

[AuCl₄]⁻ was initially deposited by electrochemical reduction on a glassy carbon electrode (GCE) to form porous nanogold layer, then prussian blue (PB) was electrodeposited onto the as-prepared nanogold layer, and then secondary nanogold particles were fabricated again on the PB surface by electrochemical reduction for the immobilization of anti-CEA antibodies. The presence of double-layer porous gold nanoparticles enhanced the immobilized amount of biomolecules, and improved the sensitivity of the immunoassay. PB, as a good redox probe, was facile to electrochemical analysis and measurement. Under optimal conditions, the developed immunoassay exhibited dynamic range from 3.0 to 80.0 ng/mL with a detection limit of 0.9 ng/mL CEA (S/N = 3). Moreover, the selectivity, reproducibility and stability of the immunosensor were acceptable.     

Resveratrol induces catalytic bioscavenger paraoxonase 1 expression and protects against chemical warfare nerve agent toxicity in human cell lines

Current advances in enzyme bioscavenger prophylactic therapy against chemical warfare nerve agent (CWNA) exposure are moving towards the identification of catalytic bioscavengers that can degrade large doses of organophosphate (OP) nerve agents without self destruction. This is a preferred method compared to therapy with the purified stoichiometric bioscavenger, butyrylcholinesterase, which binds OPs 1:1 and would thus require larger doses for treatment. Paraoxonase-1 (PON-1) is one such catalytic bioscavenger that has been shown to hydrolyze OP insecticides and contribute to detoxification in animals and humans. Here we investigated the effects of a common red wine ingredient, Resveratrol (RSV), to induce the expression of PON-1 in the human hepatic cell line HC04 and evaluated the protection against CWNA simulants. Dose-response curves showed that a concentration of 20 microM RSV was optimal in inducing PON-1 expression in HC04 cells. RSV at 20 microM increased the extracellular PON-1 activity approximately 150% without significantly affecting the cells. Higher doses of RSV were cytotoxic to the cells. Resveratrol also induced PON-1 in the human lung cell line A549. RSV pre-treatment significantly (P = 0.05) protected the hepatic cells against exposure to 2x LD(50) of soman and sarin simulants. However, lung cells were protected against soman simulant exposure but not against sarin simulant exposure following RSV treatment. In conclusion, these studies indicate that dietary inducers, such as RSV, can up-regulate PON-1, a catalytic bioscavenger, which can then hydrolyze and protect against CWNA-induced toxicity, providing a prospective new method to protect against CWNA exposure.     

Inhalation toxicity of soman vapor in non-anesthetized rats: A preliminary assessment of inhaled bronchodilator or steroid therapy

Respiratory toxicity, injury and treatment following vapor inhalational exposure to the chemical warfare nerve agent (CWNA) soman (GD) were examined in non-anesthetized rats. This study exposed male Sprague–Dawley rats (250–300 g) to 520, 560, 600, 825 or 1410 mg × min/m³ of soman in a customized head-out inhalation system. Signs of CWNA-induced cholinergic crises were observed in all soman-exposed animals. The LCt50 of vaporized soman as determined by probit analysis was 593.1 mg × min/m³. All animals exposed to 825 and 1410 mg × min/m³ developed severe convulsions and died within 4–8 min post-exposure. Edema measured by wet/dry weight ratio of the left lung lobe increased in a dose-dependent manner in all soman-exposed animals. Bronchoalveolar lavage (BAL) fluid and blood acetylcholinesterase (AChE) activities were inhibited dose-dependently in soman-exposed groups at 24 h. A significant increase in total BAL protein was observed in soman-exposed animals at all doses. AChE activity was inhibited in lung and whole brain tissues in all soman-exposed animals. Histopathological analysis of the lungs of animals exposed to 600 mg × min/m³ of soman revealed prominent morphological changes including alveolar histiocytosis, hemorrhage and inflammation consisting of neutrophilic exudate. Exposure of animals to 600 mg × min/m³ of soman followed by treatment with two actuations for 10 s of Combivent (21 μg of ipratropium bromide and 120 μg of albuterol sulfate) and Symbicort (80 μg budesonide and 4.5 μg formoterol) by inhalation into a modified metered dose inhaler (MDI) 10 min post-exposure resulted in increased minute volume, but did not decrease mortality. These results indicate that inhalation exposure to soman vapor causes acute respiratory toxicity and injury in untreated, un-anesthetized rats and that inhalation treatment with Combivent or Symbicort did improve the respiratory outcomes, but did not influence lethality.     

Role of Acetylcholinesterase on the Structure and Function of Cholinergic Synapses: Insights Gained from Studies on Knockout Mice

Electrophysiological and ultrastructural studies were performed on phrenic nerve-hemidiaphragm preparations isolated from wild-type and acetylcholinesterase (AChE) knockout (KO) mice to determine the compensatory mechanisms manifested by the neuromuscular junction to excess acetylcholine (ACh). The diaphragm was selected since it is the primary muscle of respiration, and it must adapt to allow for survival of the organism in the absence of AChE. Nerve-elicited muscle contractions, miniature endplate potentials (MEPPs) and evoked endplate potentials (EPPs) were recorded by conventional electrophysiological techniques from phrenic nerve-hemidiaphragm preparations isolated from 1.5- to 2-month-old wild-type (AChE^+/+) or AChE KO (AChE^−/−) mice. These recordings were chosen to provide a comprehensive assessment of functional alterations of the diaphragm muscle resulting from the absence of AChE. Tension measurements from AChE^−/− mice revealed that the amplitude of twitch tensions was potentiated, but tetanic tensions underwent a use-dependent decline at frequencies below 70 Hz and above 100 Hz. MEPPs recorded from hemidiaphragms of AChE^−/− mice showed a reduction in frequency and a prolongation in decay (37%) but no change in amplitude compared to values observed in age-matched wild-type littermates. In contrast, MEPPs recorded from hemidiaphragms of wild-type mice that were exposed for 30 min to the selective AChE inhibitor 5-bis(4-allyldimethyl-ammoniumphenyl)pentane-3-one (BW284C51) exhibited a pronounced increase in amplitude (42%) and a more marked prolongation in decay (76%). The difference between MEPP amplitudes and decays in AChE^−/− hemidiaphragms and in wild-type hemidiaphragms treated with BW284C51 represents effective adaptation by the former to a high ACh environment. Electron microscopic examination revealed that diaphragm muscles of AChE^−/− mice had smaller nerve terminals and diminished pre- and post-synaptic surface contacts relative to neuromuscular junctions of AChE^+/+ mice. The morphological changes are suggested to account, in part, for the ability of muscle from AChE^−/− mice to function in the complete absence of AChE.     

Neuroprotective Actions of the Synthetic Estrogen 17α-Ethynylestradiol in the Hippocampus

17α-Ethynylestradiol (EE2), a major constituent of many oral contraceptives, is similar in structure to 17β-estradiol, which has neuroprotective properties in several animal models. This study explored the potential neuroprotective actions of EE2 against kainic and quinolinic acid toxicity in the hippocampus of adult ovariectomized Wistar rats. A decrease in the number of Nissl-stained neurons and the induction of vimentin immunoreactivity in astrocytes was observed in the hilus of the dentate gyrus of the hippocampus after the administration of either kainic acid or quinolinic acid. EE2 prevented the neuronal loss and the induction of vimentin immunoreactivity induced by kainic acid at low (1 μg/rat) and high (10–100 μg/rat) doses and exerted a protection against quinolinic acid toxicity at a low dose (1 μg/rat) only. These observations demonstrate that EE2 exerts neuroprotective actions against excitotoxic insults. This finding is relevant for the design of new neuroprotective estrogenic compounds.     

The metabolic responses and acid–base status after feeding, exhaustive exercise, and both feeding and exhaustive exercise in Chinese catfish (Silurus asotus Linnaeus)

Feeding and exhaustive exercise are known to elevate metabolism. However, acid–base status may be oppositely affected by the two processes. In this study, we first investigated the acid–base response of Chinese catfish to feeding (the meal size was about 8% of body mass) to test whether an alkaline tide (a metabolic alkalosis created by gastric HCl secretion after feeding) would occur. We then determined the combined effects of feeding and exhaustive exercise on excess post-exercise oxygen consumption and acid–base status to determine whether the alkaline tide induced by feeding protects against acid–base disturbance during exhaustive exercise and affects subsequent recovery. Arterial blood pH increased from 7.74 ± 0.02 before feeding to 7.88 ± 0.02 and plasma [HCO₃ ⁻]_pl increased from 5.42 ± 0.29 to 7.83 ± 0.37 mmol L⁻¹ 6 h after feeding, while feeding had no significant effect on P_\textCO2 P_{{{\text{CO}}_{2} }} . Exhaustive exercise led to a significant reduction in pH by 0.46 units and a reduction of [HCO₃ ⁻]_pl by ~3 mmol L⁻¹. Lactate concentrations in white muscle and plasma increased by 2.4 mmol L⁻¹ and 13.4 μmol g⁻¹, respectively. Fed fish had a higher pH and [HCO₃ ⁻]_pl than fasting fish at rest, and the reductions in pH (0.36 units) and [HCO₃ ⁻]_pl (~2 mmol L⁻¹) were thus lower after exhaustive exercise. However, the recovery of acid–base status and metabolites were similar in digesting and fasting fish. Overall, a significant alkaline tide was found in Chinese catfish after feeding. The alkaline tide elicited by feeding significantly prevented the decreases in pH and [HCO₃ ⁻]_pl immediately after exhaustive exercise, but recovery from exhaustive exercise was not affected by digestion.     

Improved enzymatic production of phenolated acylglycerols through alkyl phenolate intermediates

A novel approach is reported for the synthesis of dihydrocaffoylated glycerols that consists of two steps: enzymatic synthesis of octyl dihydrocaffeate (as a synthetic intermediate) from octanol and dihydrocaffeic acid, and enzymatic interesterification of triacylglycerols with octyl dihydrocaffeate. Due to the good compatibility of the intermediate with triacylglycerols, an improved volumetric productivity [147 mol h⁻¹(kg Novozym 435)⁻¹] and high enzyme specific activity [up to 9.6 μmol⁻¹ min⁻¹(g Novozym 435)⁻¹] have been obtained.     

Inhibition of ATP-Induced Ca<Superscript>2+</Superscript> Influx by Corticosterone in Dorsal Root Ganglion Neurons

In addition to the classic genomic effects, it is well known that glucocorticoids also have rapid, nongenomic effects on neurons. In the present study, the effect of corticosterone (CORT) on ATP-induced Ca²⁺ mobilization in cultured dorsal root ganglion (DRG) neurons were detected with confocal laser scanning microscopy using fluo-4/AM as a calcium fluorescent indicator that could monitor real-time alterations of intracellular calcium concentration ([Ca²⁺]i). ATP, an algesic agent, caused [Ca²⁺]i increase in DRG neurons by activation of P2X receptor. Pretreatment with CORT (1 nM–1 μM for 5 min) inhibited ATP-induced [Ca²⁺]i increase in DRG neurons. The rapid inhibition of ATP-induced Ca²⁺ response by CORT was concentration-dependent, reversible and could be blocked by glucocorticoid receptor antagonist RU38486 (10 μM). Furthermore, the inhibitory effect of CORT was abolished by protein kinase A inhibitor H89 (10 μM), but was not influenced by protein kinase C inhibitor Chelerythrine chloride (10 μM). On the other hand, membrane-impermeable bovine serum albumin-conjugated corticosterone had no effect on ATP-induced [Ca²⁺]i transients. These observations suggest that a nongenomic pathways may be involved in the effect of CORT on ATP-induced [Ca²⁺]i transients in cultured DRG neurons.     

Relationship between reversible acetylcholinesterase inhibition and efficacy against soman lethality

Carbamate pretreatment (45% inhibition, reversible), combined with therapy, protected rats from soman-induced lethality [The Pharmacologist 23, 224 (1981)]. The present study was done to see if less than 45% inhibition protects and to see if reversible acetylcholinesterase (AChE) inhibition and efficacy against soman lethality are correlated. At 30 min pre-soman, guinea pigs and rats received (im) either pyridostigmine (Py) or physostigmine (Ph) to inhibit whole blood AChE from 10 to 70%; at 1 min post-soman (sc), they received (im) atropine (16 mg/kg)/2-PAMCl (50 mg/kg) and mecamylamine (0.8 mg/kg)/atropine (16 mg/kg), respectively. Protective ratios (PRs) were computed and they ranged from 3.1 to 7.7 for guinea pigs and from 1.8 to 2.4 for rats. In guinea pigs the PRs for Py + therapy were roughly similar to those of Ph + therapy. In both species at 30 min after im injection of Py and Ph, a linear relationship was found between percentage of whole blood AChE inhibition and ln dosage of carbamate. Positive correlation (p less than 0.05) was found between the degree of reversible AChE inhibition by pretreatment, coupled with therapy, and efficacy against soman lethality. The present data indicate that inhibition levels as low as 10% may provide some protection.     

Radioprotection by the histone deacetylase inhibitor phenylbutyrate

The histone deacetylase inhibitor (HDAC), phenylbutyrate (PB), is a novel anti-tumor agent. Studies have demonstrated that HDAC inhibitors can suppress cutaneous radiation syndrome and stimulate hematopoiesis. The objective of this study was to test the ability of PB treatment to protect against acute gamma-radiation-induced lethality in the DBA/2 mouse model. A 30-day radiation lethality study was used to assess radioprotective capability of PB. Mechanisms were evaluated using western blots, flow cytometry, and the single-cell gel electrophoresis assay. Western blot studies showed that PB treatment acetylated histones in vivo. For radiation protection studies, prophylactic administration of PB (24 h preradiation; 1–50 mg/kg) provided radioprotection against gamma radiation (8–9.5 Gy) and PB demonstrated a DRF of 1.31 (P = 0.001; 95% confidence interval: 1.27, 1.36). When PB (10 mg/kg) was administered post-radiation (4 h), it also provided significant radioprotection at 8.0 Gy radiation (P = 0.022). PB treatment before radiation was associated with significant elevations in neutrophils and platelets following radiation. Results from single-cell gel electrophoresis of peripheral blood leukocytes demonstrated that PB treatment before radiation can attenuate DNA damage and inhibit radiation-induced apoptosis. These results indicate that an HDAC inhibitor like PB has potential as a radiation protector and that mechanisms of action include attenuation of DNA damage and inhibition of apoptosis.     

New multipotent tetracyclic tacrines with neuroprotective activity

The synthesis and the biological evaluation (neuroprotection, voltage dependent calcium channel blockade, AChE/BuChE inhibitory activity and propidium binding) of new multipotent tetracyclic tacrine analogues (5–13) are described. Compounds 7, 8 and 11 showed a significant neuroprotective effect on neuroblastoma cells subjected to Ca²⁺ overload or free radical induced toxicity. These compounds are modest AChE inhibitors [the best inhibitor (11) is 50-fold less potent than tacrine], but proved to be very selective, as for most of them no BuChE inhibition was observed. In addition, the propidium displacement experiments showed that these compounds bind AChE to the peripheral anionic site (PAS) of AChE and, consequently, are potential agents that can prevent the aggregation of β-amyloid. Overall, compound 8 is a modest and selective AChE inhibitor, but an efficient neuroprotective agent against 70 mM K⁺ and 60 μM H₂O₂. Based on these results, some of these molecules can be considered as lead candidates for the further development of anti-Alzheimer drugs.     

Purinergic signalling in rat GFSHR-17 granulosa cells: an <Emphasis Type="Italic">in vitro</Emphasis> model of granulosa cells in maturing follicles

Purinergic signalling in rat GFSHR-17 granulosa cells was characterised by Ca²⁺-imaging and perforated patch-clamp. We observed a resting intracellular Ca²⁺-concentration ([Ca²⁺]_i) of 100 nM and a membrane potential of −40 mV. This was consistent with high K⁺− and Cl⁻ permeability and a high intracellular Cl⁻ concentration of 40 mM. Application of ATP for 5–15 s every 3 min induced repeated [Ca²⁺]_i increases and a 30 mV hyperpolarization. The phospholipase C inhibitor U73122 or the IP₃-receptor antagonist 2-aminoethoethyl diphenyl borate suppressed ATP responses. Further biochemical and pharmacological experiments revealed that ATP responses were related to stimulation of P2Y₂ and P2Y₄ receptors and that the [Ca²⁺]_i increase was a prerequisite for hyperpolarization. Inhibitors of Ca²⁺-activated channels or K⁺ channels did not affect the ATP-evoked responses. Conversely, inhibitors of Cl⁻ channels hyperpolarized cells to −70 mV and suppressed further ATP-evoked hyperpolarization. We propose that P2Y₂ and P2Y₄ receptors in granulosa cells modulate Cl⁻ permeability by regulating Ca²⁺-release.     

Development of the New Group of Indole-Derived Neuroprotective Drugs Affecting Oxidative Stress

Summary 1. The role of oxidative stress, and accordingly uncontrolled reactive oxygen species generation/action, have been widely documented in a number of different neuronal pathologies. However, the concept of pharmacological interventions in prevention and therapy of oxidative stress-related diseases has not found adequate application in clinical practice. This may be due to the insufficient efficacy of drugs available, their unsuitable pharmacokinetics, side effects, toxicity, etc.2. Based on stobadine, (−)-cis-2,8-dimethyl-2,3,4,4a,5,9b-hexahydro-1H-pyrido[4,3-b]indole, a well-known antioxidant, free radical scavenger, and neuroprotectant, it was attempted to develop new stobadine derivatives with improved pharmacodynamic and toxicity profiles, on applying molecular design, synthesis and adequate tests. Stobadine molecule was modified mostly by electron donating substitution on the benzene ring and by alkoxycarbonyl substitution at N-2 position. A total of >70 derivatives were prepared.3. In a mice model of head trauma, some of the new stobadine derivatives administered i.v. immediately after the trauma, significantly improved sensomotoric outcome in the animals assessed 1 h later. Accordingly, decrease in brain edema was proved histologically as well as by brain wet weight assessment.4. Putative neuroprotective action of the compounds was confirmed on rat hippocampal slices exposed to reversible 6 min hypoxia/low glucose by analysis of synaptic transmission in CA1 region neurons. Irreversible impairment of neurotransmission resulting from the hypoxia was significantly reduced by the presence of SMe1EC2, one of the new compounds, in concentration range 0.03−10.0×10⁻⁶ mol l⁻¹. Both the neuroprotective and antioxidant effect of the compound closely resembled those of stobadine, melatonin, 21-aminosteroids, alpha-phenyl-tert-butylnitrone and others, all well-established antioxidants, except the range of effective concentrations was by 1–2 orders lower in SMe1EC2.5. A remarkable antioxidant efficacy was observed in the new compounds in rat brain homogenates exposed to iron/ascorbate system by protection of lipids and creatine kinase against the oxidative impairment. A link between the neuroprotective and antioxidant/ scavenger properties in the compounds can be assumed.6. Acute toxicity of some of the new pyridoindoles was diminished compared to stobadine. That might be due to the virtually full elimination of stobadine's undesired alpha ₁-adrenolytic activity attained by appropriate modifications of its molecule.7. The new pyridoindoles extend the range of available neuroprotectants interfering with oxidative stress in neuronal tissue.