Chloroquine Restores Ganglioside Homeostasis and Improves Pathological and Behavioral Outcomes Post-stroke in the Rat.

Perturbations of ganglioside homeostasis have been observed following stroke whereby toxic simple gangliosides GM2 and GM3 accumulate, while protective complex species GM1 and GD1 are reduced. Thus, there is a need for therapeutic interventions which can prevent ganglioside dysregulation after stroke. A pharmacological intervention using chloroquine was selected for its transient lysosomotropic properties which disrupt the activity of catabolic ganglioside enzymes. Chloroquine was administered both in vitro (0.1 µM), to primary cortical neurons exposed to GM3 toxicity, and in vivo (45 mg/kg i.p.), to 3-month-old male Wistar rats that underwent a severe stroke injury. Chloroquine was administered for seven consecutive days beginning 3 days prior to the stroke injury. Gangliosides were examined using MALDI imaging mass spectrometry at 3 and 21 days after the injury, and motor deficits were examined using the ladder task. Chloroquine treatment prevented ganglioside dysregulation 3 days post-stroke and partially prevented complex ganglioside depletion 21 days post-stroke. Exogenous GM3 was found to be toxic to primary cortical neurons which was protected by chloroquine treatment. Motor deficits were prevented in the forelimbs of stroke-injured rats with chloroquine treatment and was associated with decreased inflammation, neurodegeneration, and an increase in cell survival at the site of injury. Chloroquine administration prevents ganglioside dysregulation acutely, protects against GM3 toxicity in neurons, and is associated with long-term functional and pathological improvements after stroke in the rat. Therefore, targeting lipid dysregulation using lysosomotropic agents such as chloroquine may represent a novel therapeutic avenue for stroke injuries.

Increased Expression of Simple Ganglioside Species GM2 and GM3 Detected by MALDI Imaging Mass Spectrometry in a Combined Rat Model of Aß Toxicity and Stroke.

The aging brain is often characterized by the presence of multiple comorbidities resulting in synergistic damaging effects in the brain as demonstrated through the interaction of Alzheimer's disease (AD) and stroke. Gangliosides, a family of membrane lipids enriched in the central nervous system, may have a mechanistic role in mediating the brain's response to injury as their expression is altered in a number of disease and injury states. Matrix-Assisted Laser Desorption Ionization (MALDI) Imaging Mass Spectrometry (IMS) was used to study the expression of A-series ganglioside species GD1a, GM1, GM2, and GM3 to determine alteration of their expression profiles in the presence of beta-amyloid (Aß) toxicity in addition to ischemic injury. To model a stroke, rats received a unilateral striatal injection of endothelin-1 (ET-1) (stroke alone group). To model Aß toxicity, rats received intracerebralventricular (i.c.v.) injections of the toxic 25-35 fragment of the Aß peptide (Aß alone group). To model the combination of Aß toxicity with stroke, rats received both the unilateral ET-1 injection and the bilateral icv injections of Aß25-35 (combined Aß/ET-1 group). By 3 d, a significant increase in the simple ganglioside species GM2 was observed in the ischemic brain region of rats who received a stroke (ET-1), with or without Aß. By 21 d, GM2 levels only remained elevated in the combined Aß/ET-1 group. GM3 levels however demonstrated a different pattern of expression. By 3 d GM3 was elevated in the ischemic brain region only in the combined Aß/ET-1 group. By 21 d, GM3 was elevated in the ischemic brain region in both stroke alone and Aß/ET-1 groups. Overall, results indicate that the accumulation of simple ganglioside species GM2 and GM3 may be indicative of a mechanism of interaction between AD and stroke.

Protein markers of cerebrovascular disruption of neurovascular unit: immunohistochemical and imaging approaches.

Currently, there is great interest in the assembly and function of cerebral endothelial, glial and neuronal cells that form the anatomical basis of the neurovascular unit (NVU) to maintain blood-brain barrier (BBB) and cerebral blood flow. Recent studies have provided considerable insight into the assembly of the components of the NVU. However, there is still paucity of data regarding the identification and expression pattern of these components in various pathologies of the central nervous system. Here, we provide a brief overview of the histological and imaging methods to study BBB disruption in various experimental settings especially the ischemia models associated with necrosis. Emphasis is on the immunohistochemistry of various protein markers of NVU. An understanding of the alterations in the expression pattern of these markers in various neurodegenerative disorders could lead to a better understanding of the conditions that cause BBB disruption as well as to the development of new restorative and protective treatments.