Calcium responses of chicken trigeminal ganglion neurons to methyl anthranilate and capsaicin

Using digital fluorescence imaging, we determined the effects of methyl anthranilate (MA), an avian irritant, and capsaicin (CAP), a mammalian irritant, on intracellular calcium ([Ca2+]i) in chicken trigeminal neurons. Concentration–response functions indicated that the threshold for inducing increases in [Ca2+]i was higher for CAP (30·mmol·l–1) than for MA (10·mmol·l–1). The maximum magnitudes of [Ca2+]i in response to MA and CAP were compared after normalization to 40·mmol·l–1 KCl. At equal concentrations (300·mmol·l–1), trigeminal neurons responded with a greater change in [Ca2+]i to MA (78% of KCl) than to CAP (43% of KCl). Furthermore, at 300·mmol·l–1, 48% of neurons responded to MA whereas only 16% responded to CAP. The increases in [Ca2+]i induced by both MA and CAP were dependent upon extracellular calcium. While the calcium responses to MA were also dependent on extracellular sodium, responses to CAP were not. There were separate but overlapping populations of neurons sensitive to MA and CAP. Taken together, the higher threshold concentration of CAP, the higher response magnitude to MA than CAP and the greater number of neurons sensitive to MA than CAP provide a rationale for the observed behavioral differences of birds to these two compounds. Finally, the findings that the calcium responses to MA and CAP have different ion dependencies and that there are separate populations sensitive to these compounds suggest different transduction mechanisms mediating chicken trigeminal responses to MA and CAP

Roadmap for C9ORF72 in Frontotemporal Dementia and Amyotrophic Lateral Sclerosis: Report on the C9ORF72 FTD/ALS Summit

A summit held March 2023 in Scottsdale, Arizona (USA) focused on the intronic hexanucleotide expansion in the C9ORF72 gene and its relevance in frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS; C9ORF72-FTD/ALS). The goal of this summit was to connect basic scientists, clinical researchers, drug developers, and individuals affected by C9ORF72-FTD/ALS to evaluate how collaborative efforts across the FTD-ALS disease spectrum might break down existing disease silos. Presentations and discussions covered recent discoveries in C9ORF72-FTD/ALS disease mechanisms, availability of disease biomarkers and recent advances in therapeutic development, and clinical trial design for prevention and treatment for individuals affected by C9ORF72-FTD/ALS and asymptomatic pathological expansion carriers. The C9ORF72-associated hexanucleotide repeat expansion is an important locus for both ALS and FTD. C9ORF72-FTD/ALS may be characterized by loss of function of the C9ORF72 protein and toxic gain of functions caused by both dipeptide repeat (DPR) proteins and hexanucleotide repeat RNA. C9ORF72-FTD/ALS therapeutic strategies discussed at the summit included the use of antisense oligonucleotides, adeno-associated virus (AAV)-mediated gene silencing and gene delivery, and engineered small molecules targeting RNA structures associated with the C9ORF72 expansion. Neurofilament light chain, DPR proteins, and transactive response (TAR) DNA-binding protein 43 (TDP-43)-associated molecular changes were presented as biomarker candidates. Similarly, brain imaging modalities (i.e., magnetic resonance imaging [MRI] and positron emission tomography [PET]) measuring structural, functional, and metabolic changes were discussed as important tools to monitor individuals affected with C9ORF72-FTD/ALS, at both pre-symptomatic and symptomatic disease stages. Finally, summit attendees evaluated current clinical trial designs available for FTD or ALS patients and concluded that therapeutics relevant to FTD/ALS patients, such as those specifically targeting C9ORF72, may need to be tested with composite endpoints covering clinical symptoms of both FTD and ALS. The latter will require novel clinical trial designs to be inclusive of all patient subgroups spanning the FTD/ALS spectrum

An amyloidogenic hexapeptide derived from amylin attenuates inflammation and acute lung injury in murine sepsis

<div><p>Although the accumulation of amyloidogenic proteins in neuroinflammatory conditions is generally considered pathologic, in a murine model of multiple sclerosis, amyloid-forming fibrils, comprised of hexapeptides, are anti-inflammatory. Whether these molecules modulate systemic inflammatory conditions remains unknown. We hypothesized that an amylin hexapeptide that forms fibrils can attenuate the systemic inflammatory response in a murine model of sepsis. To test this hypothesis, mice were pre-treated with either vehicle or amylin hexapeptide (20 μg) at 12 hours and 6 hours prior to intraperitoneal (i.p.) lipopolysaccharide (LPS, 20 mg/kg) administration. Illness severity and survival were monitored every 6 hours for 3 days. Levels of pro- (IL-6, TNF-α, IFN-γ) and anti-inflammatory (IL-10) cytokines were measured via ELISA at 1, 3, 6, 12, and 24 hours after LPS (i.p.). As a metric of lung injury, pulmonary artery endothelial cell (PAEC) barrier function was tested 24 hours after LPS administration by comparing lung wet-to-dry ratios, Evan’s blue dye (EBD) extravasation, lung histology and caspase-3 activity. Compared to controls, pretreatment with amylin hexapeptide significantly reduced mortality (p<0.05 at 72 h), illness severity (p<0.05), and pro-inflammatory cytokine levels, while IL-10 levels were elevated (p<0.05). Amylin pretreatment attenuated LPS-induced lung injury, as demonstrated by decreased lung water and caspase-3 activity (p<0.05, versus PBS). Hence, in a murine model of systemic inflammation, pretreatment with amylin hexapeptide reduced mortality, disease severity, and preserved lung barrier function. Amylin hexapeptide may represent a novel therapeutic tool to mitigate sepsis severity and lung injury.</p></div

Pretreatment with amylin attenuates lung injury in an LPS-induced systemic inflammatory mouse model.

<p>Adult female B6 mice, with or without amylin pretreatment, underwent induction of systemic inflammation as described prior. At 24 hours after LPS (i.p.) administration, lung tissues were inflated, fixed and evaluated for lung injury. (A) Evans Blue Dye (EBD) incorporation in lung tissue was determined by retro-orbital injection of EBD 30 min prior to the 24h experimental endpoint in PBS-only, LPS, and LPS+amylin treated mice. (B) Excessive lung water accumulation was measured using the wet/dry weight ratios of lung tissues. All values presented are mean ± SEM, with n = 10 mice/group. *p<0.05 **p<0.005 ***p<0.001 vs. PBS, via 1-way ANOVA and Bonferroni’s multiple comparisons test. Both EBD and lung water accumulation were significantly elevated in the LPS group and did not differ between PBS and LPS+AMY groups. (C) Representative H and E staining of lungs at 20X magnification from mice treated with PBS+AMY, LPS+PBS, or LPS+AMY. In animals treated with LPS, air space disease was evident with fluid in the alveoli, increased interstitial prominence, and hyaline membrane formation. In animals that received amylin pretreatment, air space disease and lung water was decreased, compared to LPS. n = 5 mice/group.</p