All known biological functions of the pro-inflammatory cytokine interleukin-1β (IL-1β) are mediated by type 1 interleukin receptor (IL-1R1). IL-1β–IL-1R1 signaling modulates various neuronal functions including spinal pain processing. Although the role of IL-1β in pain processing is generally accepted, there is a discussion in the literature whether IL-1β exerts its effect on spinal pain processing by activating neuronal or glial IL-1R1. To contribute to this debate, here we investigated the expression and cellular distribution of IL-1R1 in the superficial spinal dorsal horn in control animals and also in inflammatory pain. Experiments were performed on rats and wild type as well as IL-1R1-deficient mice. Inflammatory pain was evoked by unilateral intraplantar injection of complete Freund adjuvant (CFA). The nociceptive responsiveness of control and CFA-treated animals were tested daily for withdrawal responses to mechanical and thermal stimuli before and after CFA injection. Changes in the expression of 48 selected genes/mRNAs and in the quantity of IL-1R1 protein during the first 3 days after CFA injection were measured with the TaqMan low-density array method and Western blot analysis, respectively. The cellular localization of IL-1R1 protein was investigated with single and double staining immunocytochemical methods. We found a six times and two times increase in IL-1R1 mRNA and protein levels, respectively, in the dorsal horn of CFA-injected animals 3 days after CFA injection, at the time of the summit of mechanical and thermal allodynia. Studying the cellular distribution of IL-1R1, we found an abundant expression of IL-1R1 on the somatodendritic compartment of neurons and an enrichment of the receptor in the postsynaptic membranes of some excitatory synapses. In contrast to the robust neuronal localization, we observed only a moderate expression of IL-1R1 on astrocytes and a negligible one on microglial cells. CFA injection into the hind paw caused a remarkable increase in the expression of IL-1R1 in neurons, but did not alter the glial expression of the receptor. The results suggest that IL-1β exerts its effect on spinal pain processing primarily through neuronal IL-1R1, but it can also interact in some extent with IL-1R1 expressed by astrocytes.

The small intestine is the longest part of the chicken (Gallus gallus) gastrointestinal system that is specialized for nutrient absorption. It is known that decrease in intestinal villi area or height in early age can cause a reduction in essential nutrient intake, which may lead to delayed growth and consequently poorer performance of broiler chickens. The small intestinal absorptive surface is known to be affected by various factors, among others things the nutritional state. In our experiment, we aimed to investigate the possible protein expression alterations that lie behind the villus area and height decrease caused by feed deprivation. A total of 24 chickens were divided into three groups, namely ad libitum fed, fasted for 24 h, fasted for 24 h then refed for 2 h. The morphometric parameters were also measured in the duodenum, jejunum and ileum tissue sections using image analysis. Differential proteome analyses from jejunum samples were performed using two-dimensional difference gel electrophoresis followed by tryptic digestion and protein identification by matrix-assisted laser desorption/ionization mass spectrometry. Overall 541 protein spots were detected after 2D. Among them, eleven showed 1.5-fold or higher significant difference in expression and were successfully identified. In response to 24 h fasting, the expression of nine proteins was higher and that of two proteins was lower compared to the ad libitum fed group. The functions of the differentially expressed proteins indicate that the 24 h fasting mainly affects the expression of structural proteins, and proteins involved in lipid transport, general stress response, and intestinal defense.

The phosphorylation of serine 10 in histone 3 (p-S10H3) has recently been demonstrated to participate in spinal nociceptive processing. However, superficial dorsal horn (SDH) neurons involved in p-S10H3-mediated nociception have not been fully characterized. In the present work, we combined immunohistochemistry, in situ hybridization with the retrograde labeling of projection neurons to reveal the subset of dorsal horn neurons presenting an elevated level of p-S10H3 in response to noxious heat (60 °C), causing burn injury. Projection neurons only represented a small percentage (5%) of p-S10H3-positive cells, while the greater part of them belonged to excitatory SDH interneurons. The combined immunolabeling of p-S10H3 with markers of already established interneuronal classes of the SDH revealed that the largest subset of neurons with burn injury-induced p-S10H3 expression was dynorphin immunopositive in mice. Furthermore, the majority of p-S10H3-expressing dynorphinergic neurons proved to be excitatory, as they lacked Pax-2 and showed Lmx1b-immunopositivity. Thus, we showed that neurochemically heterogeneous SDH neurons exhibit the upregulation of p-S10H3 shortly after noxious heat-induced burn injury and consequential tissue damage, and that a dedicated subset of excitatory dynorphinergic neurons is likely a key player in the development of central sensitization via the p-S10H3 mediated pathway.

Methionine (Met) is an essential and first limiting amino acid in the poultry diet that plays a significant role in chicken embryonic development and growth. The present study examined the effect of in ovo injection of DL-Met and L-Met sources and genotypes on chicken embryonic-intestinal development and health. Fertilized eggs of the two genotypes, TETRA-SL layer hybrid (TSL) - commercial layer hybrid and Hungarian Partridge colored hen breed (HPC) - a native genotype, were randomly distributed into four treatments for each genotype. The treatment groups include the following: 1) control non-injected eggs (NoIn); 2) saline-injected (SaIn); 3) DL-Met injected (DLM); and 4) L-Met injected (LM). The in ovo injection was carried out on 17.5 d of embryonic development; after hatching, eight chicks per group were sacrificed, and the jejunum was extracted for analysis. The results showed that both DLM and LM groups had enhanced intestinal development as evidenced by increased villus width, villus height, and villus area (P < 0.05) compared to the control. The DLM group had significantly reduced crypt depth, glutathione content (GSH), glutathione S-transferase 3 alpha (GST3), occludin (OCLN) gene expression and increased villus height to crypt depth ratio in the TSL genotype than the LM group (P < 0.05). The HPC genotype has overexpressed insulin-like growth factor 1 (IGF1) gene, tricellulin (MD2), occludin (OCLN), superoxide dismutase 1 (SOD1), and GST3 genes than the TSL genotype (P < 0.05). In conclusion, these findings showed that in ovo injection of Met enhanced intestinal development, and function, with genotypes responding differently under normal conditions. Genotypes also influenced the expression of intestinal antioxidants, tight junction, and growth-related genes.

Afferents from the C2 spinal nerve (SN) and trigeminal nerve (TN) innervate neighboring cranial territories, and their convergence on the upper cervical dorsal horn neurons represents neural substrate of pain referral in primary headache disorders. Unfortunately, little is known about trigeminocervical input to the major spinal nociceptive projection area lamina I. Here, we used ex vivo brainstem–cervical cord preparation for the visually guided whole-cell recording from the upper cervical lamina I neurons. We show that 50% of them receive convergent monosynaptic input from both nerves, whereas 35% and 11% of neurons receive specific supply from the C2 SN and TN, respectively. Altogether, 10 distinct patterns of synaptic input from the C2 SN and TN to lamina I neurons could be identified. Although stimulation of both nerves evoked excitatory/inhibitory responses, more numerous pure inhibitory inputs arose from the TN. We show that cervical and trigeminal nociceptors converge on to lamina I projection and inhibitory neurons. Thus, trigeminocervical input in lamina I is processed in both nerve-specific and convergent circuitries. Afferent convergence on to inhibitory interneurons serves as a feedforward mechanism balancing excitatory drive to projection neurons. Disruption of this balance may cause pain in primary headache syndromes.

Burn injury is a trauma resulting in tissue degradation and severe pain, which is processed first by neuronal circuits in the spinal dorsal horn. We have recently shown that in mice, excitatory dynorphinergic (Pdyn) neurons play a pivotal role in the response to burn-injury-associated tissue damage via histone H3.1 phosphorylation-dependent signaling. As Pdyn neurons were mostly associated with mechanical allodynia, their involvement in thermonociception had to be further elucidated. Using a custom-made AAV9_mutH3.1 virus combined with the CRISPR/cas9 system, here we provide evidence that blocking histone H3.1 phosphorylation at position serine 10 (S10) in spinal Pdyn neurons significantly increases the thermal nociceptive threshold in mice. In contrast, neither mechanosensation nor acute chemonociception was affected by the transgenic manipulation of histone H3.1. These results suggest that blocking rapid epigenetic tagging of S10H3 in spinal Pdyn neurons alters acute thermosensation and thus explains the involvement of Pdyn cells in the immediate response to burn-injury-associated tissue damage.