Knockdown of the Non-canonical Wnt Gene Prickle2 Leads to Cerebellar Purkinje Cell Abnormalities While Cerebellar-Mediated Behaviors Remain Intact.

Autism spectrum disorders (ASD) involve brain wide abnormalities that contribute to a constellation of symptoms including behavioral inflexibility, cognitive dysfunction, learning impairments, altered social interactions, and perceptive time difficulties. Although a single genetic variation does not cause ASD, genetic variations such as one involving a non-canonical Wnt signaling gene, Prickle2, has been found in individuals with ASD. Previous work looking into phenotypes of Prickle2 knock-out (Prickle2-/-) and heterozygous mice (Prickle2-/+) suggest patterns of behavior similar to individuals with ASD including altered social interaction and behavioral inflexibility. Growing evidence implicates the cerebellum in ASD. As Prickle2 is expressed in the cerebellum, this animal model presents a unique opportunity to investigate the cerebellar contribution to autism-like phenotypes. Here, we explore cerebellar structural and physiological abnormalities in animals with Prickle2 knockdown using immunohistochemistry, whole-cell patch clamp electrophysiology, and several cerebellar-associated motor and timing tasks, including interval timing and eyeblink conditioning. Histologically, Prickle2-/- mice have significantly more empty spaces or gaps between Purkinje cells in the posterior lobules and a decreased propensity for Purkinje cells to fire action potentials. These structural cerebellar abnormalities did not impair cerebellar-associated behaviors as eyeblink conditioning and interval timing remained intact. Therefore, although Prickle-/- mice show classic phenotypes of ASD, they do not recapitulate the involvement of the adult cerebellum and may not represent the pathophysiological heterogeneity of the disorder.

PACAP Induces Light Aversion in Mice by an Inheritable Mechanism Independent of CGRP.

The neuropeptides CGRP (calcitonin gene-related peptide) and PACAP (pituitary adenylate cyclase-activating polypeptide) have emerged as mediators of migraine, yet the potential overlap of their mechanisms remains unknown. Infusion of PACAP, like CGRP, can cause migraine in people, and both peptides share similar vasodilatory and nociceptive functions. In this study, we have used light aversion in mice as a surrogate for migraine-like photophobia to compare CGRP and PACAP and ask whether CGRP or PACAP actions were dependent on each other. Similar to CGRP, PACAP induced light aversion in outbred CD-1 mice. The light aversion was accompanied by increased resting in the dark, but not anxiety in a light-independent open field assay. Unexpectedly, about one-third of the CD-1 mice did not respond to PACAP, which was not seen with CGRP. The responder and nonresponder phenotypes were stable, inheritable, and not sex linked, although there was a trend for greater responses among male mice. RNA-sequencing analysis of trigeminal ganglia yielded hierarchical clustering of responder and nonresponder mice and revealed a number of candidate genes, including greater expression of the Trpc5 and Kcnk12 ion channels and glycoprotein hormones and receptors in a subset of male responder mice. Importantly, an anti-PACAP monoclonal antibody could block PACAP-induced light aversion but not CGRP-induced light aversion. Conversely, an anti-CGRP antibody could not block PACAP-induced light aversion. Thus, we propose that CGRP and PACAP act by independent convergent pathways that cause a migraine-like symptom in mice.SIGNIFICANCE STATEMENT The relationship between the neuropeptides CGRP (calcitonin gene-related peptide) and PACAP (pituitary adenylate cyclase-activating polypeptide) in migraine is relevant given that both peptides can induce migraine in people, yet to date only drugs that target CGRP are available. Using an outbred strain of mice, we were able to show that most, but not all, mice respond to PACAP in a preclinical photophobia assay. Our finding that CGRP and PACAP monoclonal antibodies do not cross-inhibit the other peptide indicates that CGRP and PACAP actions are independent and suggests that PACAP-targeted drugs may be effective in patients who do not respond to CGRP-based therapeutics.

Amylin Analog Pramlintide Induces Migraine-like Attacks in Patients.

OBJECTIVE: Migraine is a prevalent and disabling neurological disease. Its genesis is poorly understood, and there remains unmet clinical need. We aimed to identify mechanisms and thus novel therapeutic targets for migraine using human models of migraine and translational models in animals, with emphasis on amylin, a close relative of calcitonin gene-related peptide (CGRP). METHODS: Thirty-six migraine without aura patients were enrolled in a randomized, double-blind, 2-way, crossover, positive-controlled clinical trial study to receive infusion of an amylin analogue pramlintide or human αCGRP on 2 different experimental days. Furthermore, translational studies in cells and mouse models, and rat, mouse and human tissue samples were conducted. RESULTS: Thirty patients (88%) developed headache after pramlintide infusion, compared to 33 (97%) after CGRP (p = 0.375). Fourteen patients (41%) developed migraine-like attacks after pramlintide infusion, compared to 19 patients (56%) after CGRP (p = 0.180). The pramlintide-induced migraine-like attacks had similar clinical characteristics to those induced by CGRP. There were differences between treatments in vascular parameters. Human receptor pharmacology studies showed that an amylin receptor likely mediates these pramlintide-provoked effects, rather than the canonical CGRP receptor. Supporting this, preclinical experiments investigating symptoms associated with migraine showed that amylin treatment, like CGRP, caused cutaneous hypersensitivity and light aversion in mice. INTERPRETATION: Our findings propose amylin receptor agonism as a novel contributor to migraine pathogenesis. Greater therapeutic gains could therefore be made for migraine patients through dual amylin and CGRP receptor antagonism, rather than selectively targeting the canonical CGRP receptor. ANN NEUROL 2021;89:1157-1171.

Characterization of Antibodies against Receptor Activity-Modifying Protein 1 (RAMP1): A Cautionary Tale.

Calcitonin gene-related peptide (CGRP) is a key component of migraine pathophysiology, yielding effective migraine therapeutics. CGRP receptors contain a core accessory protein subunit: receptor activity-modifying protein 1 (RAMP1). Understanding of RAMP1 expression is incomplete, partly due to the challenges in identifying specific and validated antibody tools. We profiled antibodies for immunodetection of RAMP1 using Western blotting, immunocytochemistry and immunohistochemistry, including using RAMP1 knockout mouse tissue. Most antibodies could detect RAMP1 in Western blotting and immunocytochemistry using transfected cells. Two antibodies (844, ab256575) could detect a RAMP1-like band in Western blots of rodent brain but not RAMP1 knockout mice. However, cross-reactivity with other proteins was evident for all antibodies. This cross-reactivity prevented clear conclusions about RAMP1 anatomical localization, as each antibody detected a distinct pattern of immunoreactivity in rodent brain. We cannot confidently attribute immunoreactivity produced by RAMP1 antibodies (including 844) to the presence of RAMP1 protein in immunohistochemical applications in brain tissue. RAMP1 expression in brain and other tissues therefore needs to be revisited using RAMP1 antibodies that have been comprehensively validated using multiple strategies to establish multiple lines of convincing evidence. As RAMP1 is important for other GPCR/ligand pairings, our results have broader significance beyond the CGRP field.

Patients With Vestibular Migraine are More Likely to Have Occipital Headaches than those With Migraine Without Vestibular Symptoms.

OBJECTIVE: To determine whether patients with vestibular migraine are more likely to suffer from an occipital headache than patients with migraine without vestibular symptoms. BACKGROUND: Vestibular migraine is an underdiagnosed disorder in which migraine is associated with vestibular symptoms. Anatomical evidence and symptomatology hint at the involvement of brain structures in the posterior fossa (back of the head location). We hypothesized that vestibular migraine patients are more likely than migraineurs without vestibular symptoms to experience headaches located in the back of the head, that is, occipital headaches. METHODS: A retrospective cross-sectional study was conducted at the University of Iowa Hospital and Clinics. Chart analysis of 169 patients was performed. The primary outcome was the location of the headache in vestibular migraine patients and migraineurs without vestibular symptoms. The secondary outcomes included the association of vestibular migraine with gender, age at onset of headache, age at onset of vestibular symptoms (such as vertigo, head motion-induced dizziness), aura, motion sickness, other associated symptoms, family history of headaches, and family history of motion sickness. RESULTS: In vestibular migraine group, 45/103 (44%) had occipital location for their headaches vs 12/66 (18%) in migraine patients without vestibular symptoms, for an odd's ratio of 3.5 (95% CI = 1.7-7.2, P < .001). Additionally, the age at onset of headache was greater in the vestibular migraine group (28 ± 12 vs 18 ± 9 years, P < .001) and motion sickness was more common (41/98 (42%) in the vestibular migraine group, 1/64 (2%) in the migraine without vestibular symptoms group, P < .001). CONCLUSIONS: This study suggests that patients with vestibular migraine are more likely to have occipital headaches than patients with migraine without vestibular symptoms. Our data support the initiation of a prospective study to determine whether a patient presenting with occipital headaches, with late onset of age of headache, and with a history of motion sickness is at an increased risk for the possible development of vestibular migraine.

Stimulation of Posterior Thalamic Nuclei Induces Photophobic Behavior in Mice.

OBJECTIVE: A hallmark of migraine is photophobia. In mice, photophobia-like behavior is induced by calcitonin gene-related peptide (CGRP), a neuropeptide known to be a key player in migraine. In this study, we sought to identify sites within the brain from which CGRP could induce photophobia. DESIGN: We focused on the posterior thalamic region, which contains neurons responsive to both light and dural stimulation and has CGRP binding sites. We probed this area with both optogenetic stimulation and acute CGRP injections in wild-type mice. Since the light/dark assay has historically been used to investigate anxiety-like responses in animals, we measured anxiety in a light-independent open field assay and asked if stimulation of a brain region, the periaqueductal gray, that induces anxiety would yield similar results to posterior thalamic stimulation. The hippocampus was used as an anatomical control to ensure that light-aversive behaviors could not be induced by the stimulation of any brain region. RESULTS: Optogenetic activation of neuronal cell bodies in the posterior thalamic nuclei elicited light aversion in both bright and dim light without an anxiety-like response in an open field assay. Injection of CGRP into the posterior thalamic region triggered similar light-aversive behavior without anxiety. In contrast to the posterior thalamic nuclei, optogenetic stimulation of dorsal periaqueductal gray cell bodies caused both light aversion and an anxiety-like response, while CGRP injection had no effect. In the dorsal hippocampus, neither optical stimulation nor CGRP injection affected light aversion or open field behaviors. CONCLUSION: Stimulation of posterior thalamic nuclei is able to initiate light-aversive signals in mice that may be modulated by CGRP to cause photophobia in migraine.

Seizures are regulated by ubiquitin-specific peptidase 9 X-linked (USP9X), a de-ubiquitinase.

Epilepsy is a common disabling disease with complex, multifactorial genetic and environmental etiology. The small fraction of epilepsies subject to Mendelian inheritance offers key insight into epilepsy disease mechanisms; and pathologies brought on by mutations in a single gene can point the way to generalizable therapeutic strategies. Mutations in the PRICKLE genes can cause seizures in humans, zebrafish, mice, and flies, suggesting the seizure-suppression pathway is evolutionarily conserved. This pathway has never been targeted for novel anti-seizure treatments. Here, the mammalian PRICKLE-interactome was defined, identifying prickle-interacting proteins that localize to synapses and a novel interacting partner, USP9X, a substrate-specific de-ubiquitinase. PRICKLE and USP9X interact through their carboxy-termini; and USP9X de-ubiquitinates PRICKLE, protecting it from proteasomal degradation. In forebrain neurons of mice, USP9X deficiency reduced levels of Prickle2 protein. Genetic analysis suggests the same pathway regulates Prickle-mediated seizures. The seizure phenotype was suppressed in prickle mutant flies by the small-molecule USP9X inhibitor, Degrasyn/WP1130, or by reducing the dose of fat facets a USP9X orthologue. USP9X mutations were identified by resequencing a cohort of patients with epileptic encephalopathy, one patient harbored a de novo missense mutation and another a novel coding mutation. Both USP9X variants were outside the PRICKLE-interacting domain. These findings demonstrate that USP9X inhibition can suppress prickle-mediated seizure activity, and that USP9X variants may predispose to seizures. These studies point to a new target for anti-seizure therapy and illustrate the translational power of studying diseases in species across the evolutionary spectrum.

The bed nucleus of the stria terminalis is critical for anxiety-related behavior evoked by CO2 and acidosis.

Carbon dioxide (CO2) inhalation lowers brain pH and induces anxiety, fear, and panic responses in humans. In mice, CO2 produces freezing and avoidance behavior that has been suggested to depend on the amygdala. However, a recent study in humans with bilateral amygdala lesions revealed that CO2 can trigger fear and panic even in the absence of amygdalae, suggesting the importance of extra-amygdalar brain structures. Because the bed nucleus of the stria terminalis (BNST) contributes to fear- and anxiety-related behaviors and expresses acid-sensing ion channel-1A (ASIC1A), we hypothesized that the BNST plays an important role in CO2-evoked fear-related behaviors in mice. We found that BNST lesions decreased both CO2-evoked freezing and CO2-conditioned place avoidance. In addition, we found that CO2 inhalation caused BNST acidosis and that acidosis was sufficient to depolarize BNST neurons and induce freezing behavior; both responses depended on ASIC1A. Finally, disrupting Asic1a specifically in the BNST reduced CO2-evoked freezing, whereas virus-vector-mediated expression of ASIC1A in the BNST of Asic1a(-/-) and Asic1a(+/+) mice increased CO2-evoked freezing. Together, these findings identify the BNST as an extra-amygdalar fear circuit structure important in CO2-evoked fear-related behavior.

Defective motile cilia in Prickle2-deficient mice.

Motile cilia play diverse roles across phyla and cell types, and abnormalities in motile cilia lead to numerous disease states, including hydrocephalus. Although motile ciliary abnormalities in Prickle2 mutants have not yet been described, the planar cell polarity genes, including Prickle2, are implicated in the development and function of motile cilia. This report evaluates Prickle2-deficient mice for dysfunction in processes known to depend on functioning motile cilia. Prickle2-deficient mice do not develop hydrocephalus, but do display abnormal morphology and motility in the motile cilia of the ependyma. The morphology of tracheal motile cilia is also abnormal. Taken together, these results demonstrate that Prickle2 is required for normal ependymal motile cilia development and function.

Duckweed application in nature-based system for water phytoremediation and high-value coproducts at family agrisystem from a circular economy perspective.

Duckweeds are widely recognized for their efficiency in the phytoremediation of agricultural and industrial effluents. This study had two main objectives: 1) Implement a Nature-based Solutions (NBS) utilizing the environmental services of duckweeds to improve water quality through phytoremediation in small fish farms; 2) Analysis of duckweeds biomass produced in these fish farms to develop coproducts from a circular economy perspective in family agrisystem in Brazilian Atlantic Forest. The effectiveness of the phytoremediation system was assessed by the reduction of the Trophic State Index (TSI). Phytoremediation in small fish farming NBS was implemented using Clarias gariepinus, employing two different managements approaches: (i) System I - L. minor cultured every 15 days, with biomass harvest and effluent analysis conducted in each cycle over 60 days; (ii) System II - L. minor cultured every 30 days, following a similar cycle and analysis. Additionally, effluent from fish production underwent testing for phytoremediation in a batch system within a climate-controlled laboratory. L. minor demonstrated efficiency in System II, leading to a reduction of the TSI. The dry biomass of the plants emerged as a viable source of amino acid for application in functional foods and feed or nutraceuticals. The findings underscore the potential integration of L. minor into the NBS system and the generation of new co-products from circular production. In addition to its effective phyto- remediation properties, L. minor's dry biomass exhibited appealing characteristics, with elevated levels of crude protein, minerals, fatty acids, and carotenoids. This positions L. minor as a promising candidate for developing bioproducts tailored for functional foods and nutraceuticals. This underscores the potential of duckweeds to produce valuable nutritional compounds beyond their remediation capabilities.

Observation of nitrogen vacancy photoluminescence from an optically levitated nanodiamond.

We present what we believe to be the first evidence of nitrogen vacancy (NV) photoluminescence (PL) from a nanodiamond suspended in a free-space optical dipole trap at atmospheric pressure. The PL rates are shown to decrease with increasing trap laser power, but are inconsistent with a thermal quenching process. For a continuous-wave trap, the neutral charge state (NV(0)) appears to be suppressed. Chopping the trap laser yields higher total count rates and results in a mixture of both NV(0) and the negative charge state (NV(-).

Evaluation of root surface microtopography following the use of four instrumentation systems by confocal microscopy and scanning electron microscopy: an in vitro study.

BACKGROUND AND OBJECTIVE: The ideal instrument for initial periodontal therapy should enable the removal of all extraneous substances from the root surfaces without any iatrogenic effects. Because of that the objective of this study is to analyse and to compare the root surface roughness after using Gracey curettes, termination diamond burs (40 µm), a piezo-ceramic ultrasonic scaler and a piezosurgery ultrasonic scaler using confocal microscopy and scanning electron microscopy. MATERIAL AND METHODS: A 2 mm × 2 mm interproximal root area of 20 teeth (n = 40 surfaces) was evaluated by confocal microscopy (×20 magnification) and scanning electron microscopy (×50 to ×1000 magnification). Teeth were randomly assigned to the following four groups: Gracey curettes with 15 vertical strokes; termination diamond burs (40 µm) at 3000 r.p.m.; a piezo-ceramic ultrasonic scaler with a power of 11; and a piezosurgery ultrasonic scaler in mode ROOT with a power of two. RESULTS: Confocal microscopy revealed that curettes [mean changes in the value of surface roughness average reduced by 0.11 ± 0.3], piezo-ceramic ultrasonic scaler (roughness average reduced by 0.47 ± 0.93) and piezosurgery ultrasonic scaler (roughness average reduced by 0.62 ± 0.93) left a smoother surface than termination diamond burs (roughness average increased by 0.39 ± 0.18). Statistically significant differences were observed in roughness (p = 0.005) between piezosurgery and termination diamond burs (p = 0.005). No statistically significant differences were between piezosurgery and Gracey curettes (p = 0.140) and between piezosurgery and piezo-ceramic ultrasonic scalers (p = 0.745). Confocal microscopy and scanning electron microscopy showed that piezosurgery seems to leave the smoothest surface. Surfaces treated with termination burs appear to show more scratches and pits. CONCLUSION: Three of the four instruments tested for root planing reduced surface roughness; however, the piezosurgery ultrasonic scaler produced the smoothest surface. The termination diamond burs (40 µm) produced a rougher surface than the ultrasonic instruments and the hand curettes. Further clinical studies are needed.

CGRP Administration Into the Cerebellum Evokes Light Aversion, Tactile Hypersensitivity, and Nociceptive Squint in Mice.

The neuropeptide calcitonin gene-related peptide (CGRP) is a major player in migraine pathophysiology. Previous preclinical studies demonstrated that intracerebroventricular administration of CGRP caused migraine-like behaviors in mice, but the sites of action in the brain remain unidentified. The cerebellum has the most CGRP binding sites in the central nervous system and is increasingly recognized as both a sensory and motor integration center. The objective of this study was to test whether the cerebellum, particularly the medial cerebellar nuclei (MN), might be a site of CGRP action. In this study, CGRP was directly injected into the right MN of C57BL/6J mice via a cannula. A battery of tests was done to assess preclinical behaviors that are surrogates of migraine-like symptoms. CGRP caused light aversion measured as decreased time in the light zone even with dim light. The mice also spent more time resting in the dark zone, but not the light, along with decreased rearing and transitions between zones. These behaviors were similar for both sexes. Moreover, significant responses to CGRP were seen in the open field assay, von Frey test, and automated squint assay, indicating anxiety, tactile hypersensitivity, and spontaneous pain, respectively. Interestingly, CGRP injection caused significant anxiety and spontaneous pain responses only in female mice, and a more robust tactile hypersensitivity in female mice. No detectable effect of CGRP on gait was observed in either sex. These results suggest that CGRP injection in the MN causes light aversion accompanied by increased anxiety, tactile hypersensitivity, and spontaneous pain. A caveat is that we cannot exclude contributions from other cerebellar regions in addition to the MN due to diffusion of the injected peptide. These results reveal the cerebellum as a new site of CGRP actions that may contribute to migraine-like hypersensitivity.

Stimulation of CGRP-expressing neurons in the medial cerebellar nucleus induces light and touch sensitivity in mice.

Calcitonin gene-related peptide (CGRP) is considered a major player in migraine pathophysiology. However, the location and mechanisms of CGRP actions in migraine are not clearly elucidated. One important question yet to be answered is: Does central CGRP signaling play a role in migraine? One candidate site is the cerebellum, which serves as a sensory and motor integration center and is activated in migraine patients. The cerebellum has the most CGRP binding sites in the central nervous system and a deep cerebellar nucleus, the medial nucleus (MN), expresses CGRP (MNCGRP). A previous study demonstrated that CGRP delivery into the cerebellum induced migraine-like behaviors. We hypothesized that stimulation of MNCGRP neurons might induce migraine-like behaviors. To test the hypothesis, we used an optogenetic strategy using CalcaCre/+ mice to drive Cre-dependent expression of channelrhodopsin-2 selectively in CGRP neurons in the cerebellar MN. A battery of behavioral tests was done to assess preclinical behaviors that are surrogates of migraine symptoms, including light aversion, cutaneous allodynia, and spontaneous pain when MNCGRP neurons were optically stimulated. Motor functions were also assessed. Optical stimulation of MNCGRP neurons decreased the time spent in the light, which was coupled to increased time spent resting in the dark, but not the light. These changes were only significant in female mice. Plantar tactile sensitivity was increased in the ipsilateral paws of both sexes, but contralateral paw data were less clear. There was no significant increase in anxiety-like behavior, spontaneous pain (squint), or changes in gait. These discoveries reveal that MNCGRP neurons may contribute to migraine-like sensory hypersensitivity to light and touch.

Involvement of the cerebellum in migraine.

Migraine is a disabling neurological disease characterized by moderate or severe headaches and accompanied by sensory abnormalities, e.g., photophobia, allodynia, and vertigo. It affects approximately 15% of people worldwide. Despite advancements in current migraine therapeutics, mechanisms underlying migraine remain elusive. Within the central nervous system, studies have hinted that the cerebellum may play an important sensory integrative role in migraine. More specifically, the cerebellum has been proposed to modulate pain processing, and imaging studies have revealed cerebellar alterations in migraine patients. This review aims to summarize the clinical and preclinical studies that link the cerebellum to migraine. We will first discuss cerebellar roles in pain modulation, including cerebellar neuronal connections with pain-related brain regions. Next, we will review cerebellar symptoms and cerebellar imaging data in migraine patients. Lastly, we will highlight the possible roles of the neuropeptide calcitonin gene-related peptide (CGRP) in migraine symptoms, including preclinical cerebellar studies in animal models of migraine.

Automated detection of squint as a sensitive assay of sex-dependent calcitonin gene-related peptide and amylin-induced pain in mice.

ABSTRACT: We developed an automated squint assay using both black C57BL/6J and white CD1 mice to measure the interpalpebral fissure area between the upper and lower eyelids as an objective quantification of pain. The automated software detected a squint response to the commonly used nociceptive stimulus formalin in C57BL/6J mice. After this validation, we used the automated assay to detect a dose-dependent squint response to a migraine trigger, the neuropeptide calcitonin gene-related peptide, including a response in female mice at a dose below detection by the manual grimace scale. Finally, we found that the calcitonin gene-related peptide amylin induced squinting behavior in female mice, but not males. These data demonstrate that an automated squint assay can be used as an objective, real-time, continuous-scale measure of pain that provides higher precision and real-time analysis compared with manual grimace assessments.

Investigating Migraine-Like Behavior Using Light Aversion in Mice.

Migraine is a complex neurological disorder characterized by headache and sensory abnormalities, such as hypersensitivity to light, observed as photophobia. Whilst it is impossible to confirm that a mouse is experiencing migraine, light aversion can be used as a behavioral surrogate for the migraine symptom of photophobia. To test for light aversion, we utilize the light/dark assay to measure the time mice freely choose to spend in either a light or dark environment. The assay has been refined by introducing two critical modifications: pre-exposures to the chamber prior to running the test procedure and adjustable chamber lighting, permitting the use of a range of light intensities from 55 lux to 27,000 lux. Because the choice to spend more time in the dark is also indicative of anxiety, we also utilize a light-independent anxiety test, the open field assay, to distinguish anxiety from light-aversive behavior. Here, we describe a modified test paradigm for the light/dark and open field assays. The application of these assays is described for intraperitoneal injection of calcitonin gene-related peptide (CGRP) in two mouse strains and for optogenetic brain stimulation studies.

Contrast-enhanced proton radiographic sensitivity limits for tumor detection.

Purpose: Proton radiography may guide proton therapy cancer treatments with beam's-eye-view anatomical images and a proton-based estimation of proton stopping power. However, without contrast enhancement, proton radiography will not be able to distinguish tumor from tissue. To provide this contrast, functionalized, high- Z nanoparticles that specifically target a tumor could be injected into a patient before imaging. We conducted this study to understand the ability of gold, as a high- Z , biologically compatible tracer, to differentiate tumors from surrounding tissue. Approach: Acrylic and gold phantoms simulate a tumor tagged with gold nanoparticles (AuNPs). Calculations correlate a given thickness of gold to levels of tumor AuNP uptake reported in the literature. An identity, × 3 , and × 7 proton magnifying lens acquired lens-refocused proton radiographs at the 800-MeV LANSCE proton beam. The effects of gold in the phantoms, in terms of percent density change, were observed as changes in measured transmission. Variable areal densities of acrylic modeled the thickness of the human body. Results: A 1 - µ m -thick gold strip was discernible within 1 cm of acrylic, an areal density change of 0.2%. Behind 20 cm of acrylic, a 40 - µ m gold strip was visible. A 1-cm-diameter tumor tagged with 1 × 10 5 50-nm AuNPs per cell has an amount of contrast agent embedded within it that is equivalent to a 65 - µ m thickness of gold, an areal density change of 0.63% in a tissue thickness of 20 cm, which is expected to be visible in a typical proton radiograph. Conclusions: We indicate that AuNP-enhanced proton radiography might be a feasible technology to provide image-guidance to proton therapy, potentially reducing off-target effects and sparing nearby tissue. These data can be used to develop treatment plans and clinical applications can be derived from the simulations.

Different forms of traumatic brain injuries cause different tactile hypersensitivity profiles.

ABSTRACT: Chronic complications of traumatic brain injury represent one of the greatest financial burdens and sources of suffering in the society today. A substantial number of these patients suffer from posttraumatic headache (PTH), which is typically associated with tactile allodynia. Unfortunately, this phenomenon has been understudied, in large part because of the lack of well-characterized laboratory animal models. We have addressed this gap in the field by characterizing the tactile sensory profile of 2 nonpenetrating models of PTH. We show that multimodal traumatic brain injury, administered by a jet-flow overpressure chamber that delivers a severe compressive impulse accompanied by a variable shock front and acceleration-deceleration insult, produces long-term tactile hypersensitivity and widespread sensitization. These are phenotypes reminiscent of PTH in patients, in both cephalic and extracephalic regions. By contrast, closed head injury induces only transient cephalic tactile hypersensitivity, with no extracephalic consequences. Both models show a more severe phenotype with repetitive daily injury for 3 days, compared with either 1 or 3 successive injuries in a single day, providing new insight into patterns of injury that may place patients at a greater risk of developing PTH. After recovery from transient cephalic tactile hypersensitivity, mice subjected to closed head injury demonstrate persistent hypersensitivity to established migraine triggers, including calcitonin gene-related peptide and sodium nitroprusside, a nitric oxide donor. Our results offer the field new tools for studying PTH and preclinical support for a pathophysiologic role of calcitonin gene-related peptide in this condition.

Calcitonin gene-related peptide (CGRP): role in migraine pathophysiology and therapeutic targeting.

Introduction: The neuropeptide calcitonin gene-related peptide (CGRP) is recognized as a critical player in migraine pathophysiology. Excitement has grown regarding CGRP because of the development and clinical testing of drugs targeting CGRP or its receptor. While these drugs alleviate migraine symptoms in half of the patients, the remaining unresponsive half of this population creates an impetus to address unanswered questions that exist in this field.Areas covered: We describe the role of CGRP in migraine pathophysiology and CGRP-targeted therapeutics currently under development and in use. We also discuss how a second CGRP receptor may provide a new therapeutic target.Expert opinion: CGRP-targeting drugs have shown a remarkable safety profile. We speculate that this may reflect the redundancy of peptides within the CGRP family and a second CGRP receptor that may compensate for reduced CGRP activity. Furthermore, we propose that an inherent safety feature of peptide-blocking antibodies is attributed to the fundamental nature of peptide release, which occurs as a large bolus in short bursts of volume transmission. These facts support the development of more refined CGRP therapeutic drugs, as well as drugs that target other neuropeptides. We believe that the future of migraine research is bright with exciting advances on the horizon.