Full-length ACTH molecule and ACTH-related hexapeptide H-PKKRRP-OH are demonstrated by the patch-clamp method to decrease the NaV1.8 channel activation gating system effective charge in the nociceptive neuron membrane, while ACTH-related tetrapeptide Ac-KKRR-NH2 has no effect. ACTH(1-24), a fully functional ACTH mimetic, and H-PKKRRP-OH show analgesic effects in the formalin test in vivo. All peptides contain the cationic KKRR motif, but only H-PKKRRP-OH and ACTH(1-24) relieve acute pain, targeting the NaV1.8 channel as a receptor. This seemingly controversial result is explained by application of conformational analysis and blind docking. Though conformational analysis indicates that both H-PKKRRP-OH and Ac-KKRR-NH2 contain the cationic functional groups at the earlier suggested characteristic distance of 9-12 Å, Ac-KKRR-NH2 does not interact with the S4I voltage sensor of the NaV1.8 channel activation gating system. The docking demonstrates that an extensive network of ligand-receptor ionic and hydrogen bonds involving D151, E157, R218, and R221 VSDI residues, essential for the analgesic tripeptide Ac-KKK-NH2 binding, is formed upon the H-PKKRRP-OH binding. Particularly important are the ionic bonds between the H-PKKRRP-OH C-terminal carboxylate anion and the S4I R218 and R221 guanidinium groups. The described mechanism of NaV1.8 channel modulation is fundamentally different from the effect of channel blockers.

A potential analgesic medicinal substance has been discovered, the ouabain–Ca2+ chelate complex (EO). As we have found, the specific EO binding to the Na,K-ATPase (NKA) in nanomolar concentrations triggers several signaling cascades in the nociceptive neuron, two of which have been discussed elsewhere. The docking results indicate that the molecular basis for the specificity of EO–NKA binding is the formation of two intermolecular ionic bonds between the chelated Ca2+ cation and two NKA carboxylate anion, Glu116 and Glu117. The third downstream EO-triggered NKA/Src/PKA/p38 MAPK/NF-κB signaling pathway, likely, controls the GAP43 gene expression, which results in this case in the neurite-inhibiting effect at the tissue level. The strong EO analgesic effect at both the spinal and supraspinal levels has been demonstrated in the formalin test. EO is a promising candidate for the role of a novel and safe analgesic, which might be particularly effective for the treatment of the tumor-associated pain syndromes due to its possible cytostatic function.

Two short arginine-containing tripeptides, H-Arg-Arg-Arg-OH (TP1) and Ac-Arg-Arg-Arg-NH2 (TP2), have been shown by the patch-clamp method to modulate the NaV1.8 channels of DRG primary sensory neurons, which are responsible for the generation of nociceptive signals. Conformational analysis of the tripeptides indicates that the key role in the ligand-receptor binding of TP1 and TP2 to the NaV1.8 channel is played by two positively charged guanidinium groups of the arginine side chains located at the characteristic distance of ~9 Å from each other. The tripeptide effect on the NaV1.8 channel activation gating device has been retained when the N- and C-terminal groups of TP1 were structurally modified to TP2 to protect the attacking peptide from proteolytic cleavage by exopeptidases during its delivery to the molecular target, the NaV1.8 channel. As demonstrated by the organotypic tissue culture method, the agents do not affect the DRG neurite growth, which makes it possible to expect the absence of adverse side effects at the tissue level upon administration of TP1 and TP2. The data obtained indicate that both tripeptides can have great therapeutic potential as novel analgesic medicinal substances.

Spatial learning, memory, and reactivity of the hypothalamic—pituitary—adrenocortical system (HPA axis) were studied in adult male rats, whose mothers during pregnancy were subjected to acute moderate normobaric hypoxia, or repeated injections of buspirone, an agonist of type 1A serotonergic receptors (5HT1A), or their combination. Prenatal treatment with buspirone in rats with prenatal hypoxia impaired learning ability during the first day of 5-day training. A decrease in the effectiveness of long-term memory in comparison with short-term memory was revealed in two groups of rats: prenatal treatment with buspirone in combination with hypoxia and injection of physiological saline without hypoxia. The effectiveness of long-term memory and the level of corticosterone in response to stress did not differ between the groups, which can indicate adaptation of the 5HT1A receptor and the HPA axis to the prenatal buspirone and normobaric hypoxia during ontogeny.

To address the problem of neonatal pain-related stress effects on spatial learning, memory and the stress reactivity of the hypothalamo–pituitary–adrenal axis (the HPA) in adolescent and adult rats we gave male and female rats an intraplantar injection of moderate concentration of formalin (2.5 0.5 µL) (Formalin rats) on postnatal days (P1 and P2). Adolescent male Formalin rats showed expressed impairments in both learning and memory. Adult male and female Formalin rats displayed better spatial learning than the adolescent Formalin rats; males, regardless of age, revealed better ability in learning than females. Adult Formalin males showed higher HPA axis reactivity, which was combined with a higher performance of long-term memory compared with the corresponding data in adult females. The data demonstrate that the consequences of moderate neonatal pain-related stress manifested themselves in later life differently depending on the age and sex. Moreover, the results testify to an unconventional adaptive potential of pain-related stress, which, when interacting with behavioral plasticity, improved the functioning of neurobiological systems in adult rats. The identified sex differences in effects of pain-related stress on the cognitive functions in rats of both ages, as well as the stress reactivity of hormonal response in adult rats, suggest sexual dimorphism in neonatal pain-triggered synaptic plasticity of structures involved in spatial learning and memory.

The present work continues our recent series of articles that aim to elucidate the ligand–receptor binding mechanism of short cationic peptides to the NaV1.8 channel in the nociceptive neuron. The applied methodological approach has involved several methods: the patch-clamp experimental evaluation of the effective charge of the NaV1.8 channel activation gating system, the organotypic tissue culture method, the formalin test, and theoretical conformational analysis. The lysine-containing short peptide Ac-KEKK-NH2 has been shown to effectively modulate the NaV1.8 channel activation gating system. As demonstrated by the organotypic tissue culture method, the studied short peptide does not trigger the downstream signaling cascades controlling neurite outgrowth and should not be expected to evoke adverse side effects. Conformational analysis of the Ac-KEKK-NH2 molecule has revealed that the distances between the positively charged amino groups of the lysine side chains are equal to 11–12 Å. According to the previously suggested mechanism of ligand–receptor binding of short peptides to the NaV1.8 channel molecule, Ac-KEKK-NH2 should exhibit an analgesic effect, which has been confirmed by the formalin test. The data obtained unequivocally indicate that the studied lysine-containing short peptide is a promising candidate for the role of a novel analgesic medicinal substance.

We report here a study examining the effects of acute normobaric hypoxia in the early postnatal period of development (postnatal day 2) on learning ability and the expression of GABAB receptors in the neocortex (medial prefrontal cortex) of adolescent rats (postnatal days 55–60) and also addressing the possibility of correcting identified disorders with the Russian-made drug salifen, a GABA derivative. Perinatal hypoxia has been shown to impair the formation and retention of memory traces in the novel object recognition and conditioned passive avoidance reflex tests and to reduce spatial learning ability in the Morris water maze test. Analysis of Western blot results revealed an increase in the quantity of GABAB receptor protein in the medial prefrontal area of the neocortex of rats exposed to perinatal hypoxia. Administration of salifen 15 mg/kg for 14 days after exposure to hypoxia improved the cognitive abilities of rats and normalized the level of GABAB receptor protein expression in the medial prefrontal cortex of rats. The results obtained here are of interest for solving an applied challenge in neonatology – the search for effective pharmacological correction of the sequelae of perinatal hypoxia.

The study investigated the effects of 1) acute normobaric hypoxia with a reduced oxygen concentration in inhaled air and 2) the selective serotonin reuptake inhibitor fluoxetine in newborn female rat pups on spatial learning, the performance of spatial short-term and long-term memory and reactivity of the hypothalamic-pituitary-adrenal axis (the HPA axis) in adult rats. The newly obtained data indicate a deterioration in the ability for spatial learning only in the first out of five training days. Hypoxia did not change the performance of memory in the test “recognition of a new object” and caused an improvement in long-term memory in the Morris water maze. This was combined with a higher content of corticosterone in blood plasma after testing long-term memory. Fluoxetine did not change the performance indicators of memory or hormonal indicators in either control or hypoxic adult rats. Fluoxetine normalized the latent period of reaching the platform in hypoxic rats when testing learning, and did not reduce the indicator of long-term memory improved by hypoxia. The obtained results suggest that neonatal normobaric hypoxia under certain conditions helps improving memory, while fluoxetine has a protective effect on cognitive abilities and the HPA axis in adult female rats.

We studied the effect of moderate neonatal normobaric hypoxia on the indicators of spatial learning, memory, and reactivity of the hypothalamic-pituitary-adrenocortical system in adult male Wistar rats. The pharmacological effect of chronic injections of the serotonin reuptake inhibitor fluoxetine during the neonatal period on the studied behavioral and the physiological indices was evaluated. Hypoxia impaired spatial training, increased the short-term memory performance, but did not change long-term memory and stress indicator in response to its testing. The use of fluoxetine normalized learning, but did not change memory indicators and the stress-induced level of corticosterone in blood plasma in the hypoxic rats and control animals. New results indicate a protective effect of fluoxetine in the neonatal period under conditions of moderate normobaric hypoxia.

The effect of moderate neonatal stress induced by inflammatory pain in rat pups of both sexes on the hormonal response and cognitive processes in adult animals was studied in the Morris water maze. No significant differences in spatial learning and memory were found in experimental rats exposed to neonatal inflammatory pain vs. control animals. However, experimental rats exhibited sex differences in long-term spatial memory whose efficiency was higher in males vs. females. After long-term memory testing, stress responsiveness of the hypothalamic-pituitary-adrenocortical axis, as assessed by the plasma corticosterone level in the formalin test, was higher in experimental males vs. females. Only experimental females exhibited differences between short-term and long-term memory, with the efficiency being higher in the former. Thus, sexual dimorphism was found in the effect of neonatal nociceptive stress on long-term spatial memory in adult rats: experimental males vs. females demonstrated more effective long-term memory combined with a higher stress reactivity of the hormonal response.

Repetitive neonatal pain may cause central nervous system disorders. Existing clinical data on the effect of neonatal pain on learning, memory and stress-reactivity of the hypothalamic-pituitary-adrenal axis (the HPA axis) are limited to adolescence and have mostly been obtained for male individuals. The mechanism of neonatal pain effect has not yet been researched. Ontogenetic studies carried out on different-sex specimen are crucial for prediction, prevention and treatment of adaptive behavioural disorders caused by neonatal pain stress. Lately, this has become even more relevant due to COVID-19 affecting new-borns in need of intensive therapy. In this study, the impact of neonatal inflammatory pain on adult cognition and hormonal stress response was investigated in male and female rats. Continuous neonatal pain was induced by hindpaw formalin injections on days one and two from birth. In adult rats, we assessed spatial learning and memory using the Morris water maze and evaluated HPA reactivity in response to the formalin test. No significant cognitive function differences were found between the experimental group of rats subjected to neonatal pain and the control group. Within the experimental group subjected to neonatal pain, males showed better long-term spatial memory than females. After testing the long-term memory, we evaluated HPA reactivity by corticosterone levels in response to a formalin test. This parameter was higher in males. Only females with neonatal pain showed differences between short-term and long-term memory, with poorer long-term memory. Possible causes of sex differences in the cognitive function and hormonal stress response in adult rats exposed to neonatal inflammatory pain are discussed.

We investigated the effect of neonatal inflammatory pain onthe dynamics of plasma corticosterone level in newborn rat pupsand, in the late prepubertal period (days 45–53), the processesof spatial learning and memory, as well as the reactivity of thehypothalamic–pituitary–adrenal (HPA) axis to forced swim stress,in male and female rats. We found a long-term increase in HPA axisreactivity in response to peripheral inflammatory pain induced byformalin injection to newborn rat pups, which suggested the developmentalmodification of the HPA axis and its possible involvement in theregulation of cognitive abilities. Neonatal pain caused a deficitin spatial learning and memory, revealing a sexual dimorphism inthe implementation of these processes. After recording long-termmemory in 53-day-old rats in the forced swim test, the plasma corticosteronewas found to be increased compared to the basal level, however,no differences in the hormone level were detected both in rats withneonatal pain vs. control animals and in males vs. females. Thus,in the late prepubertal period, rats showed no direct relationshipbetween neonatal pain-induced activation of the HPA axis and theindices of spatial learning and memory, which may indicate the involvementof other neurobiological systems in the effect of neonatal painon cognitive abilities.

Exposure to stress at an early age programs the HPA axis which can lead to cognitive deficits in adults. However, it is not known whether these deficits emerge in adulthood or are expressed earlier in life. The aims of the study were to investigate (1) the immediate effects of early injury-induced stress in one-day-old (P1) and repeated stress on at P1 and P2 rat pups on plasma corticosterone levels; and (2) examine the subsequent long-term effects of this early stress on spatial learning and memory, and stress reactivity in early P26-34 and late P45-53 adolescent male and female rats. Intra-plantar injection of formalin induced prolonged and elevated levels of corticosterone in pups and impaired spatial learning and short- and long-term memory in late adolescent males and long-term memory in early adolescent females. There were sex differences in late adolescence in both learning and short-term memory. Performance on the long-term memory task was better than that on the short-term memory task for all early adolescent male and female control and stressed animals. Short-term memory was better in the late age control rats of both sexes and for formalin treated females as compared with the early age rats. These results are consistent with an impaired function of structures involved in memory (the hippocampus, amygdala, prefrontal cortex) after newborn pain. However, activation of the HPA axis by neonatal pain did not directly correlate with spatial learning and memory outcomes and the consequences of neonatal pain remain are likely multi-determined.

The aim of the work was to study pathomorphological changes in the testes in males with babesiosis. Diagnosis of babesiosis in sick males was performed based on the detection of typical clinical signs and blood smears. Blood smears were fixed in methanol and stained according to the Romanovsky-Giemsa method with hematoxylin and eosin. For histological studies, pieces of testicles were selected, which were fixed in an 8% neutral aqueous formalin solution. The prepared material was poured into paraffin according to the conventional method. Histological sections were stained with hematoxylin and eosin using the method of Van Gieson and Mallory. Pathomorphological examination of stained histological sections revealed changes characteristic of parenchymal orchitis. The spermatogenic epithelium of the convoluted seminal tubules was drained down in all fields of vision. The lumen of the tubules was filled with protein detritus, exfoliated cells of the spermatogenic epithelium and macrophages. Part of the Sertoli cells was in a state of vacuolar degeneration. Focal clusters of lymphoid-macrophage infiltrates were visible between the convoluted seminal tubules, especially multiple ones around the blood vessels. Endocrine cells were subjected to atrophy due to compression by cell infiltrates and connective tissue growths.