Construction of a Global Pain Systems Network Highlights Phospholipid Signaling as a Regulator of Heat Nociception

The ability to perceive noxious stimuli is critical for an animal's survival in the face of environmental danger, and thus pain perception is likely to be under stringent evolutionary pressure. Using a neuronal-specific RNAi knock-down strategy in adult Drosophila, we recently completed a genome-wide functional annotation of heat nociception that allowed us to identify α2δ3 as a novel pain gene. Here we report construction of an evolutionary-conserved, system-level, global molecular pain network map. Our systems map is markedly enriched for multiple genes associated with human pain and predicts a plethora of novel candidate pain pathways. One central node of this pain network is phospholipid signaling, which has been implicated before in pain processing. To further investigate the role of phospholipid signaling in mammalian heat pain perception, we analysed the phenotype of PIP5Kα and PI3Kγ mutant mice. Intriguingly, both of these mice exhibit pronounced hypersensitivity to noxious heat and capsaicin-induced pain, which directly mapped through PI3Kγ kinase-dead knock-in mice to PI3Kγ lipid kinase activity. Using single primary sensory neuron recording, PI3Kγ function was mechanistically linked to a negative regulation of TRPV1 channel transduction. Our data provide a systems map for heat nociception and reinforces the extraordinary conservation of molecular mechanisms of nociception across different species. © 2012 Neely et al

Construction of a Global Pain Systems Network Highlights Phospholipid Signaling as a Regulator of Heat Nociception

The ability to perceive noxious stimuli is critical for an animal's survival in the face of environmental danger, and thus pain perception is likely to be under stringent evolutionary pressure. Using a neuronal-specific RNAi knock-down strategy in adult Drosophila, we recently completed a genome-wide functional annotation of heat nociception that allowed us to identify α2δ3 as a novel pain gene. Here we report construction of an evolutionary-conserved, system-level, global molecular pain network map. Our systems map is markedly enriched for multiple genes associated with human pain and predicts a plethora of novel candidate pain pathways. One central node of this pain network is phospholipid signaling, which has been implicated before in pain processing. To further investigate the role of phospholipid signaling in mammalian heat pain perception, we analysed the phenotype of PIP5Kα and PI3Kγ mutant mice. Intriguingly, both of these mice exhibit pronounced hypersensitivity to noxious heat and capsaicin-induced pain, which directly mapped through PI3Kγ kinase-dead knock-in mice to PI3Kγ lipid kinase activity. Using single primary sensory neuron recording, PI3Kγ function was mechanistically linked to a negative regulation of TRPV1 channel transduction. Our data provide a systems map for heat nociception and reinforces the extraordinary conservation of molecular mechanisms of nociception across different species

Nucleus accumbens functional connectivity predicts medication overuse headache

Medication-overuse headache (MOH) is a secondary form of headache related to the overuse of acute anti-headaches. A number of studies has suggested that MOH might share the same pathophysiological and behavioral mechanisms that intervene in substance abuse and behavioral addictions (Calabresi Cupini 2008, Ferraro, Grazzi, Muffatti, Nava et al., 2012; Gómez-Beldarrain M, Carrasco M, Bilbao A, García-Moncó, 2011; Radat F, Lanteri-Minet, 2011; Calabresi, Cupini, 2005). Here, we investigated if functional resting state could be used to discriminate MOH from non-MOH migraneurs. For this aim, we used a novel methodological approach, the connectivity-based classification, to test the hypothesis that functional connectivity between the nucleus accumbens, one key structure for reward, motivational and incentive learning networks, and other cortical regions could discriminate the two forms of migraine. Methods We recorded resting state fMRI of two groups of migraine patients, age and education matched, with or without MOH. We explored the whole brain functional connectivity of the nucleus accumbens using seed-voxel correlation (Margulies et al., 2007). We then applied a pattern classification technique to investigate if resting state NAcc functional connectivity is able to discriminate between MOH and non-MOH patients. To inspect the pattern of connections having the highest discriminative power, we employed Multivoxel Pattern Analysis (MVPA). In this multivariate technique, multivoxel patterns of foci were analyzed using a method that combines machine learning with an iterative, multivariate voxel selection algorithm: Recursive Feature Elimination (RFE) (De Martino et al., ) Results Seed-voxel correlations of the resting state activity in the nucleus accumbens were found with the insula bilaterally, orbitofrontal and cingulate cortices, thalamus, basal ganglia, amydgala. We also detected connections between the accumbens and sensorimotor cortices (see Figure 2). Connectivity based classification MVPA approach demonstrated that the two groups can be efficiently discriminated on the basis of the NAcc resting state functional connectivity. Areas that showed the highest discriminative power for MOH patients were prevalently placed in the bilateral anterior and posterior insulae, dorsolateral prefrontal cortices, midcingulate cortices, precuneus, secondary sensorymotor cortices and thalami; Areas that showed the highest discriminative power for non-MOH patients were prevalently placed in sensorimotor and premotor brain areas. To better interpret the findings, we grouped MVPA results into large scale brain networks. This visualization approach showed that discrimination patterns that predict abusers (MOH patients) are prevalently in orbitofrontal cortex (OFC), dorsal attention network (DAN), default model network (DMN), saliency network (SAL), cerebellar, and sensorimotor networks. Conversely, non-abusers (non-MOH patients) were predicted by the ventral attention network (VAN), motor and premotor networks. Conclusion Our data show that MOH patients are characterized by functional alteration of motivational circuits at rest. These peculiarities are sufficiently evident to allow a blind discrimination of MOH and non-MOH patients. Altogether these findings provide important suggestions in the development of novel and objective diagnostic measures

Intravenous Thrombolysis for Acute Ischemic Stroke Associated to Extracranial Internal Carotid Artery Occlusion: The ICARO-2 Study

Background and Purposes: In a case-control study in patients with acute ischemic stroke and extracranial internal carotid artery (eICA) occlusion, thrombolytic treatment was associated with increased mortality. The aim of this cohort study was to assess the efficacy and safety of thrombolysis in patients with eICA occlusion compared to those without eICA occlusion. Methods: Consecutive patients treated with intravenous tissue-type plasminogen activator within 4.5 h from symptom onset included in the Safe Implementation of Thrombolysis in Stroke - International Stroke Thrombolysis Registry (SITS-ISTR) in 20 Italian centres were analyzed. Acute carotid occlusion was diagnosed using ultrasound examination, angio-CT scan or angio-MRI. Since the SITS-ISTR database did not plan to report the site of vessel occlusion, each participating center provided the code of the patient with eICA occlusion. Patients were divided into 2 groups, those with and those without eICA occlusion. Main outcome measures were: death, disability (modified Rankin Scale, mRS, 3-6) and any intracranial bleeding at 3 months. Multiple logistic regression analysis was performed to reveal predictors for main outcomes. The following variables of interest were included in the analysis: presence of eICA occlusion, age, gender, diabetes mellitus, hyperlipidemia, atrial fibrillation, congestive heart failure, previous stroke, current smoking, antiplatelet treatment at stroke onset, baseline NIHSS score, baseline blood glucose, cholesterol and blood pressure, history of hypertension and stroke onset to treatment time. Results: A total of 1,761 patients without eICA occlusion and 137 with eICA occlusion were included in the study. At 3 months, 42 patients were lost to follow-up (3 with eICA occlusion). Death occurred in 30 (22.4%) patients with eICA occlusion and in 175 (10.2%) patients without (p < 0.0001). Death or disability at 3 months occurred in 91 of 134 patients with eICA occlusion (67.9%) compared with 654 of 1,722 patients without eICA occlusion (37.9%, p < 0.0001). No or minimal disability at 3 months (mRS 0-1) was reported in 25 (18.7%) patients with eICA occlusion and in 829 (48.2%) patients without (p < 0.0001). Any intracranial bleeding detected by CT or MRI at posttreatment imaging was seen in 16 (11.7%) patients with eICA occlusion and in 314 (17.8%) of those without (p = 0.09). The proportion of symptomatic intracerebral hemorrhage was 5.8% for patients with eICA occlusion and 8.0% for patients without (p = 0.16). At logistic regression analysis, eICA occlusion was associated with mortality (odds ratio, OR 5.7; 95% confidence interval, CI 2.9-11.1) and mortality or disability (OR 5.0; 95% CI 2.9-8.7) at 90 days. Conclusions: This cohort study in patients with acute ischemic stroke treated with thrombolysis showed an association between eICA occlusion and adverse outcome. Copyright (C) 2012 S. Karger AG, Base

Intravenous Thrombolysis for Acute Ischemic Stroke Associated to Extracranial Internal Carotid Artery Occlusion: The ICARO-2 Study

BACKGROUND AND PURPOSES: In a case-control study in patients with acute ischemic stroke and extracranial internal carotid artery (eICA) occlusion, thrombolytic treatment was associated with increased mortality. The aim of this cohort study was to assess the efficacy and safety of thrombolysis in patients with eICA occlusion compared to those without eICA occlusion. METHODS: Consecutive patients treated with intravenous tissue-type plasminogen activator within 4.5 h from symptom onset included in the Safe Implementation of Thrombolysis in Stroke - International Stroke Thrombolysis Registry (SITS-ISTR) in 20 Italian centres were analyzed. Acute carotid occlusion was diagnosed using ultrasound examination, angio-CT scan or angio-MRI. Since the SITS-ISTR database did not plan to report the site of vessel occlusion, each participating center provided the code of the patient with eICA occlusion. Patients were divided into 2 groups, those with and those without eICA occlusion. Main outcome measures were: death, disability (modified Rankin Scale, mRS, 3-6) and any intracranial bleeding at 3 months. Multiple logistic regression analysis was performed to reveal predictors for main outcomes. The following variables of interest were included in the analysis: presence of eICA occlusion, age, gender, diabetes mellitus, hyperlipidemia, atrial fibrillation, congestive heart failure, previous stroke, current smoking, antiplatelet treatment at stroke onset, baseline NIHSS score, baseline blood glucose, cholesterol and blood pressure, history of hypertension and stroke onset to treatment time. RESULTS: A total of 1,761 patients without eICA occlusion and 137 with eICA occlusion were included in the study. At 3 months, 42 patients were lost to follow-up (3 with eICA occlusion). Death occurred in 30 (22.4%) patients with eICA occlusion and in 175 (10.2%) patients without (p < 0.0001). Death or disability at 3 months occurred in 91 of 134 patients with eICA occlusion (67.9%) compared with 654 of 1,722 patients without eICA occlusion (37.9%, p < 0.0001). No or minimal disability at 3 months (mRS 0-1) was reported in 25 (18.7%) patients with eICA occlusion and in 829 (48.2%) patients without (p < 0.0001). Any intracranial bleeding detected by CT or MRI at posttreatment imaging was seen in 16 (11.7%) patients with eICA occlusion and in 314 (17.8%) of those without (p = 0.09). The proportion of symptomatic intracerebral hemorrhage was 5.8% for patients with eICA occlusion and 8.0% for patients without (p = 0.16). At logistic regression analysis, eICA occlusion was associated with mortality (odds ratio, OR 5.7; 95% confidence interval, CI 2.9-11.1) and mortality or disability (OR 5.0; 95% CI 2.9-8.7) at 90 days. CONCLUSIONS: This cohort study in patients with acute ischemic stroke treated with thrombolysis showed an association between eICA occlusion and adverse outcome

PI3Kγ acts in DRG neurons as a negative regulator of thermal and TRPV1 responses.

<p>(A) Representative temperature response ramps and Arrhenius plots for heat-activated currents measured in single DRG neurons isolated from wild type (WT) and <i>PI3Kγ</i> mutant (KO) mice. For temperature response ramps, red lines depict temperature ramps and black lines depict inward current. (B) Q10 as a measure of the rate of inward current changes in response to temperature. n = 37 for isolated WT; n = 9 for <i>PI3Kγ</i> KO DRG neurons. (C,D) Capsaicin sensitivity of DRG neurons isolated from WT and <i>PI3Kγ</i> KO mice. (C) Representative capsaicin responses from a single DRG neuron. (D) Dose-response curves to different concentrations of capsaicin. The capsaicin EC50 is indicated. Numbers indicate numbers of single neurons tested with the indicated capsaicin doses at the respective data points. Electrophysiology data was generated by single neuron patch clamping. Data are presented as mean +/− sem. ** p<0.01, *** p<0.001 (Mann-Whitney u-test).</p

A global network map of thermal nociception.

<p>The systems network includes data from significantly enriched <i>Drosophila</i> KEGG pathways and GO processes, mouse and human KEGG pathways and C2 gene sets. Pathways, processes and gene sets that share a role in a biological process were pooled into functional classes while the underlying genes that constitute them are depicted with a connection to their respective functional class. Functional classes (gold), genes representing direct hits with a thermal nociception phenotype (red), their first degree binding partners (green), and developmental lethal genes (blue) represent the nodes in the network. Only select KEGG pathways, biological processes and C2 gene sets were used to build systems map. For the entire list of individual pathways, gene sets, and processes see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003071#pgen.1003071.s010" target="_blank">Tables S5</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003071#pgen.1003071.s011" target="_blank">S6</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003071#pgen.1003071.s012" target="_blank">S7</a>.</p

PI5Kα signaling controls thermal and capsaicin nociception <i>in vivo</i>.

<p>(A) Thermal pain thresholds of wild type (WT) and <i>PIP5Kα</i>^−/− (KO) littermates in response to radiant heat (Hargreaves; n = 6 for WT; n = 6 for KO mice). (B) <i>PIP5Kα</i> KO mice also show enhanced thermal sensitivity in the hot plate assay (n = 12 for WT; n = 12 for KO mice) and (C) an exaggerated capsaicin-evoked behavioral response (n = 12 for WT; n = 9 for KO mice). (D) PIP5Kα KO mice exhibited normal mechanical pain (force threshold latency) as assessed by the von Frey test (n = 9 for WT; n = 9 for KO mice). All data are presented as mean +/− sem. *<i>p</i><0.05; **<i>p</i><0.01 (t-test).</p

Intravenous Thrombolysis for Acute Ischemic Stroke Associated to Extracranial Internal Carotid Artery Occlusion: The ICARO-2 Study

BACKGROUND AND PURPOSES: In a case-control study in patients with acute ischemic stroke and extracranial internal carotid artery (eICA) occlusion, thrombolytic treatment was associated with increased mortality. The aim of this cohort study was to assess the efficacy and safety of thrombolysis in patients with eICA occlusion compared to those without eICA occlusion. METHODS: Consecutive patients treated with intravenous tissue-type plasminogen activator within 4.5 h from symptom onset included in the Safe Implementation of Thrombolysis in Stroke - International Stroke Thrombolysis Registry (SITS-ISTR) in 20 Italian centres were analyzed. Acute carotid occlusion was diagnosed using ultrasound examination, angio-CT scan or angio-MRI. Since the SITS-ISTR database did not plan to report the site of vessel occlusion, each participating center provided the code of the patient with eICA occlusion. Patients were divided into 2 groups, those with and those without eICA occlusion. Main outcome measures were: death, disability (modified Rankin Scale, mRS, 3-6) and any intracranial bleeding at 3 months. Multiple logistic regression analysis was performed to reveal predictors for main outcomes. The following variables of interest were included in the analysis: presence of eICA occlusion, age, gender, diabetes mellitus, hyperlipidemia, atrial fibrillation, congestive heart failure, previous stroke, current smoking, antiplatelet treatment at stroke onset, baseline NIHSS score, baseline blood glucose, cholesterol and blood pressure, history of hypertension and stroke onset to treatment time. RESULTS: A total of 1,761 patients without eICA occlusion and 137 with eICA occlusion were included in the study. At 3 months, 42 patients were lost to follow-up (3 with eICA occlusion). Death occurred in 30 (22.4%) patients with eICA occlusion and in 175 (10.2%) patients without (p < 0.0001). Death or disability at 3 months occurred in 91 of 134 patients with eICA occlusion (67.9%) compared with 654 of 1,722 patients without eICA occlusion (37.9%, p < 0.0001). No or minimal disability at 3 months (mRS 0-1) was reported in 25 (18.7%) patients with eICA occlusion and in 829 (48.2%) patients without (p < 0.0001). Any intracranial bleeding detected by CT or MRI at posttreatment imaging was seen in 16 (11.7%) patients with eICA occlusion and in 314 (17.8%) of those without (p = 0.09). The proportion of symptomatic intracerebral hemorrhage was 5.8% for patients with eICA occlusion and 8.0% for patients without (p = 0.16). At logistic regression analysis, eICA occlusion was associated with mortality (odds ratio, OR 5.7; 95% confidence interval, CI 2.9-11.1) and mortality or disability (OR 5.0; 95% CI 2.9-8.7) at 90 days. CONCLUSIONS: This cohort study in patients with acute ischemic stroke treated with thrombolysis showed an association between eICA occlusion and adverse outcome