Optimizing intraluminal monofilament model of ischemic stroke in middle-aged Sprague-Dawley rats.

Intraluminal monofilament model of middle cerebral artery occlusion (MCAO) is widely adopted for ischemic stroke; and Sprague-Dawley (SD) rats are commonly used rodents for preclinical research. Due to the paucity of information on the appropriate monofilament size for inducing MCAO in SD rats and the importance of including middle-aged models in ischemic stroke studies, we aimed to: (i). determine an appropriate Doccol® monofilament size for middle-aged male SD rats which weighed > 500 g following 24-h transient MCAO survival as well as (ii). demonstrate the optimal Doccol® filament size for middle-aged males (≤ 500 g) and females (273-300 g) while using young adult male SD rats (372-472 g) as control for severity of infarct volume following 7-days post-MCAO. All rats were subjected to 90-min transient MCAO. We show that 0.43 mm Doccol® monofilament size is more appropriate to induce large infarct lesion and optimal functional deficit when compared to 0.45 mm and 0.47 mm at 24 h post-MCAO. Our data on infarct volumes at 7 days post-MCAO as well as the observed weight loss and functional deficits at post-MCAO days 1, 3 and 7 demonstrate that 0.41 mm, 0.37 mm and 0.39 mm are optimal Doccol® filament sizes for middle-aged male (477.3 ± 39.61 g) and female (302.6 ± 26.28 g) as well as young-adult male (362.2 ± 28.38 g) SD rats, respectively.

Sleep apnea and ischemic stroke- a perspective for translational preclinical modelling.

Obstructive sleep apnea (OSA) is associated with ischemic stroke. There is, however, a lack of knowledge on the exact cause-effect relationship, and preclinical models of OSA for experimental ischemic stroke investigations are not well characterized. In this review, we discuss sleep apnea and its relationship with stroke risk factors. We consider how OSA may lead to ischemic stroke and how OSA-induced metabolic syndrome and hypothalamic-pituitary axis (HPA) dysfunction could serve as therapeutic targets to prevent ischemic stroke. Further, we examine the translational potential of established preclinical models of OSA. We conclude that metabolic syndrome and HPA dysfunction, which are often overlooked in the context of experimental stroke and OSA studies, are crucial for experimental consideration to improve the body of knowledge as well as the translational potential of investigative efforts.

Graded reduction of Pafah1b1 (Lis1) activity results in neuronal migration defects and early embryonic lethality.

Heterozygous mutation or deletion of the beta subunit of platelet-activating factor acetylhydrolase (PAFAH1B1, also known as LIS1) in humans is associated with type I lissencephaly, a severe developmental brain disorder thought to result from abnormal neuronal migration. To further understand the function of PAFAH1B1, we produced three different mutant alleles in mouse Pafah1b1. Homozygous null mice die early in embryogenesis soon after implantation. Mice with one inactive allele display cortical, hippocampal and olfactory bulb disorganization resulting from delayed neuronal migration by a cell-autonomous neuronal pathway. Mice with further reduction of Pafah1b1 activity display more severe brain disorganization as well as cerebellar defects. Our results demonstrate an essential, dosage-sensitive neuronal-specific role for Pafah1b1 in neuronal migration throughout the brain, and an essential role in early embryonic development. The phenotypes observed are distinct from those of other mouse mutants with neuronal migration defects, suggesting that Pafah1b1 participates in a novel pathway for neuronal migration.

Platelet-activating factor produces neuronal growth cone collapse.

It is generally believed that neuronal growth cone collapsing factors are proteins that interact with membrane-bound receptors. Platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine; PAF) - a phospholipid autocoid, also interacts with a membrane-bound neuronal receptor which is similar in nature to collapsing factor receptors. We report that PAF and the nonhydrolyzable PAF agonist, methyl carbamyl PAF (1-O-hexadecyl-2N-methylcarbamyl-sn-glycero-3-phosphocholine, mc-PAF), evoke a dose-dependent neuronal growth cone collapse. This collapse is specifically attenuated by the PAF receptor antagonist BN-52021. These data point to a PAF receptor-mediated growth cone collapse. Therefore, PAF must be added to the list of collapsing factors which potentially guide axons to their proper targets in the developing nervous system.

Elvax as a slow-release delivery agent for a platelet-activating factor receptor agonist and antagonist.

The ability of the slow-release polymer, Elvax, to deliver a large number of different molecules to distinct brain regions has been well-documented. However, there are currently no reports of Elvax-mediated delivery of ether phospholipids, whose lipid structure renders them difficult to solubilize and detect under experimental conditions. In order to potentially examine the role of platelet-activating factor (PAF) receptors in the brain in vivo, we tested the ability of Elvax to release the ether phospholipid. PAF receptor agonist, methyl-carbamyl platelet-activating factor (mc-PAF), and the ginkolide, PAF receptor antagonist, BN 52021. Mc-PAF and BN 52021 containing Elvax sections (200 microns) prepared in methylene chloride were assayed for release characteristics with a rabbit platelet aggregation bioassay. We measured a slow, sustained release of mc-PAF and BN 52021 from Elvax in vitro over a 5 day period. Similar in vivo mc-PAF and BN 52021 release kinetics were determined. Therefore, we believe that this novel method of slow-release delivery of PAF receptor agonists and antagonists as measured with platelet aggregation is viable and offers a simple, sensitive and inexpensive method for potential in vivo studies of the roles of PAF and its receptors in neural systems.

Vascular neural network phenotypic transformation after traumatic injury: potential role in long-term sequelae.

The classical neurovascular unit (NVU), composed primarily of endothelium, astrocytes, and neurons, could be expanded to include smooth muscle and perivascular nerves present in both the up- and downstream feeding blood vessels (arteries and veins). The extended NVU, which can be defined as the vascular neural network (VNN), may represent a new physiological unit to consider for therapeutic development in stroke, traumatic brain injury, and other brain disorders (Zhang et al., Nat Rev Neurol 8(12):711-716, 2012). This review is focused on traumatic brain injury and resultant post-traumatic changes in cerebral blood flow, smooth muscle cells, matrix, blood-brain barrier structures and function, and the association of these changes with cognitive outcomes as described in clinical and experimental reports. We suggest that studies characterizing TBI outcomes should increase their focus on changes to the VNN, as this may yield meaningful therapeutic targets to resolve posttraumatic dysfunction.

Platelet-activating factor receptor stimulation disrupts neuronal migration In vitro.

LIS-1 is a gene whose hemi-deletion causes the human neuronal migration disorder Miller-Dieker lissencephaly. It encodes a subunit of a brain platelet-activating factor (PAF) acetylhydrolase, an enzyme that inactivates PAF by hydrolyzing the acetyl moiety in the sn2 position of this phospholipid. Because PAF receptor activation has been shown to affect the developing neuronal cytoskeleton, we have hypothesized that a role for PAF in neurodevelopment is that of a modulator of neuroblast movement (a cytoskeletal function) and that an aberrant regulation of PAF could lead to an early arrest in migration. This report examines the effects of the nonhydrolyzable PAF receptor agonist methyl carbamyl PAF (mc-PAF) on the unidirectional in vitro migration of granule cells from cerebellar cell reaggregates on a laminin substrate. Bath treatment with mc-PAF yields a dose-dependent decrease in granule cell migration compared with controls. This effect can be blocked by the simultaneous bath application of BN 52021 and trans-BTD, PAF receptor-specific antagonists. Although mc-PAF minimally inhibited neurite growth, its primary effect was on somal movement along preextended neurites. These experiments suggest that the stimulation of neuronal PAF receptors could be one crucial step for the regulation of neuroblast migration and that disturbed PAF catabolism during neurodevelopment could contribute to the neuronal migration defects observed in Miller-Dieker lissencephaly.

Neuronal platelet-activating factor receptor signal transduction involves a pertussis toxin-sensitive G-protein.

In most nonneural systems, platelet-activating factor (PAF) receptor effects are mediated by G-proteins that are often pertussis toxin-sensitive. The activation of pertussis toxin-sensitive G-proteins linked to PAF receptors results in the mobilization of intracellular calcium, at least in part, through the second messenger inositol triphosphate. We have sought to determine if a pertussis toxin-sensitive G-protein is involved in the PAF receptor-mediated phenomena of growth cone collapse and of synaptic enhancement in primary neuronal culture. Using infrared differential interference contrast microscopy and patch-clamp recording techniques, pertussis toxin, but not the inactive B oligomer of the toxin, was found to block both the growth cone collapse and the enhanced synaptic release of excitatory transmitter induced by a nonhydrolyzable PAF receptor agonist, making it likely that Go, Gq, or Gi is the G-protein transducer of PAF receptors in primary neurons. We believe that PAF acts directly on neuronal receptors, which are linked to pertussis toxin-sensitive G-proteins, on the tips of developing neurites, and on presynaptic nerve terminals, leading to growth cone collapse and enhanced synaptic release of transmitter.