The cerebrovascular endothelium exerts a profound influence on cerebral vessels and cerebral blood flow. This review summarizes current knowledge of various dilator and constrictor mechanisms intrinsic to the cerebrovascular endothelium. The endothelium contributes to the resting tone of cerebral arteries and arterioles by tonically releasing nitric oxide (NO*). Dilations can occur by stimulated release of NO*, endothelium-derived hyperpolarization factor, or prostanoids. During pathological conditions, the dilator influence of the endothelium can turn to that of constriction by a variety of mechanisms, including decreased NO* bioavailability and release of endothelin-1. The endothelium may participate in neurovascular coupling by conducting local dilations to upstream arteries. Further study of the cerebrovascular endothelium is critical for understanding the pathogenesis of a number of pathological conditions, including stroke, traumatic brain injury, and subarachnoid hemorrhage.

Retrograde viral transport (i.e., muscle to motoneuron) enables targeted gene delivery to specific motor pools. Recombinant adeno-associated virus serotype 9 (AAV9) robustly infects motoneurons, but the retrograde transport capabilities of AAV9 have not been systematically evaluated. Accordingly, we evaluated the retrograde transduction efficiency of AAV9 after direct tongue injection in 129SVE mice as well as a mouse model that displays neuromuscular pathology (Gaa -/ -). Hypoglossal (XII) motoneurons were histologically evaluated 8 weeks after tongue injection with AAV9 encoding green fluorescent protein (GFP) with expression driven by the chicken b-actin promoter (1 · 10 11 vector genomes). On average, GFP expression was detected in 234 -43 XII motoneurons 8 weeks after AAV9-GFP tongue injection. In contrast, tongue injection with a highly efficient retrograde anatomical tracer (cholera toxin b subunit, CT-b) resulted in infection of 818 -88 XII motoneurons per mouse. The retrograde transduction efficiency of AAV9 was similar between the 129SVE mice and those with neuromuscular disease (Gaa -/ -). Routine hematoxylin and eosin staining and cluster of differentiation (CD) immunostaining for T cells (CD3) indicated no persistent inflammation within the tongue or XII nucleus after AAV9 injection. Additional experiments indicated no adverse effects of AAV9 on the pattern of breathing. We conclude that AAV9 can retrogradely infect a significant portion of a given motoneuron pool in normal and dystrophic mice, and that its transduction efficiency is approximately 30% of what can be achieved with CT-b.

Pompe disease is an inherited disorder due to a mutation in the gene that encodes acid α-glucosidase (GAA). Children with infantile-onset Pompe disease develop progressive hypotonic weakness and cardiopulmonary insufficiency that may eventually require mechanical ventilation (MV). Our team conducted a first in human trial of diaphragmatic gene therapy (AAV1-CMV-GAA) to treat respiratory neural dysfunction in infantile-onset Pompe. Subjects (aged 2-15 years, full-time MV: n=5, partial/no MV: n=4) underwent a period of preoperative inspiratory muscle conditioning exercise. The change in respiratory function after exercise alone was compared to the change in function after intramuscular delivery of AAV1-CMV-GAA to the diaphragm with continued exercise. Since AAV-mediated gene therapy can reach phrenic motoneurons via retrograde transduction, we hypothesized that AAV1-CMV-GAA would improve dynamic respiratory motor function to a greater degree than exercise alone. Dependent measures were maximal inspiratory pressure (MIP), respiratory responses to inspiratory threshold loads (load compensation: LC), and physical evidence of diaphragm activity (descent on MRI, EMG activity). Exercise alone did not change function. After AAV1-CMV-GAA, MIP was unchanged. Flow and volume LC responses increased after dosing (p<0.05 to p<0.005), but only in the subjects with partial/no MV use. Changes in LC tended to occur on or after 180 days. At Day 180, the four subjects with MRI evidence of diaphragm descent had greater maximal voluntary ventilation (p<0.05) and tended to be younger, stronger, and use fewer hours of daily MV. In conclusion, combined AAV1-CMV-GAA and exercise training conferred benefits to dynamic motor function of the diaphragm. Children with a higher baseline neuromuscular function may have greater potential for functional gains.

. Effects of carbon monoxide and heme oxygenase inhibitors in cerebral vessels of rats and mice. Am J Physiol Heart Circ Physiol 291: H223-H230, 2006. First published February 17, 2006 doi:10.1152/ajpheart.00058.2006 has been postulated to be a signaling molecule in many tissues, including the vasculature. We examined vasomotor responses of adult rat and mouse cerebral arteries to both exogenously applied and endogenously produced CO. The diameter of isolated, pressurized, and perfused rat middle cerebral arteries (MCAs) was not altered by authentic CO (10 Ϫ6 to 10 Ϫ4 M). Mouse MCAs, however, dilated by 21 Ϯ 10% at 10 Ϫ4 M CO. Authentic nitric oxide (NO⅐, 10 Ϫ10 to 10 Ϫ7 M) dilated both rat and mouse MCAs. At 10 Ϫ8 M NO⅐, rat vessels dilated by 84 Ϯ 4%, and at 10 Ϫ7 M NO⅐, mouse vessels dilated by 59 Ϯ 9%. Stimulation of endogenous CO production through heme oxygenase (HO) with the heme precursor ␦-aminolevulinic acid (10 Ϫ10 to 10 Ϫ4 M) did not dilate the MCAs of either species. The metalloporphyrin HO inhibitor chromium mesoporphyrin IX (CrMP) caused profound constriction of the rat MCA (44 Ϯ 2% at 3 ϫ 10 Ϫ5 M). Importantly, this constriction was unaltered by exogenous CO (10 Ϫ4 M) or CO plus 10 Ϫ5 M biliverdine (both HO products). In contrast, exogenous CO (10 Ϫ4 M) reversed CrMP-induced constriction in rat gracilis arterioles. Control mouse MCAs constricted by only 3 Ϯ 1% in response to 10 Ϫ5 M CrMP. Magnesium protoporphyrin IX (10 Ϫ5 M), a weak HO inhibitor used to control for nonspecific effects of metalloporphyrins, also constricted the rat MCA to a similar extent as CrMP. We conclude that, at physiological concentrations, CO is not a dilator of adult rodent cerebral arteries and that metalloporphyrin HO inhibitors have nonspecific constrictor effects in rat cerebral arteries. cerebral arteries; chromium mesoporphyrin; endothelium-derived hyperpolarization factor; gracilis arteriole HEME OXYGENASE (HO) is the major enzyme responsible for degrading heme (29). Three isoforms of HO (HO-1, -2, and -3) catalyze the conversion of heme to carbon monoxide (CO), biliverdine IX␣, and iron (29,35,54). CO, in turn, has been postulated to be a signaling molecule in many tissues, including the vasculature (22, 29, 40 -42, 48, 54, 55, 58). HO-1 is not normally expressed in most tissues, although it may be upregulated by a wide variety of stimuli, including subarachnoid hemorrhage (45) and ischemia-reperfusion (38). HO-2 is constitutively expressed and is found in most tissues, including the brain, vascular smooth muscle, and endothelium (8,29,30,38,47,57). HO-3 may be constitutively expressed in the rat, although recent work suggests that it is a splice variant of HO-2 transcripts (16,31,54).CO binds to heme moieties such as those found in soluble guanylyl cyclase, nitric oxide (NO⅐) synthase, and cytochrome P-450 enzymes. Binding of CO to heme moieties can either increase or decrease enzymatic activity (7, 17, 22, 29, 40 -43, 46). Large-conductance calcium-dependent K ϩ channels can also be activated by CO (18,24,28,...

Objectives Small series have suggested that outcomes after abusive head trauma (AHT) are less favorable than after other injury mechanisms. We sought to determine the impact of AHT on mortality and identify factors that differentiate children with AHT from those with TBI from other mechanisms. Design First 200 subjects from the Approaches and Decisions in Acute Pediatric TBI (ADAPT) Trial – a comparative effectiveness study using an observational, cohort study design. Setting Pediatric intensive care units in tertiary children’s hospitals in USA and abroad. Participants Consecutive children (age <18 y) with severe TBI (GCS ≤ 8; intracranial pressure (ICP) monitoring). Interventions None Measurements and Main Results Demographics, injury-related scores, prehospital and resuscitation events were analyzed. Children were dichotomized based on likelihood of AHT. A total of 190 children were included (n = 35 with AHT). AHT subjects were younger (1.87 y ± 0.32 vs. 9.23 y ± 0.39, p < 0.001) and a greater proportion were female (54.3% vs. 34.8%, p = 0.032). AHT were more likely to (i) be transported from home (60.0% vs. 33.5%, p < 0.001), (ii) have apnea (34.3% vs. 12.3%, p = 0.002) and (iii) seizures (28.6% vs. 7.7%, p < 0.001) during pre-hospital care. AHT had a higher incidence of seizures during resuscitation (31.4 vs. 9.7%, p = 0.002). After adjusting for covariates, there was no difference in mortality (AHT, 25.7% vs. non-AHT, 18.7%, HR 1.758, p = 0.60). A similar proportion died due to refractory intracranial hypertension in each group (AHT, 66.7% vs. non-AHT, 69.0%). Conclusion In this large, multicenter series, children with AHT had differences in prehospital and in-hospital secondary injuries which could have therapeutic implications. Unlike other TBI populations in children, female predominance was seen in AHT in our cohort. Similar mortality rates and refractory ICP deaths suggest that children with severe AHT may benefit from therapies including invasive monitoring and adherence to evidenced-based guidelines.