Early treatment with minocycline following stroke in rats improves functional recovery and differentially modifies responses of peri-infarct microglia and astrocytes.

BACKGROUND: Altered neuronal connectivity in peri-infarct tissue is an important contributor to both the spontaneous recovery of neurological function that commonly develops after stroke and improvements in recovery that have been induced by experimental treatments in animal models. Microglia and astrocytes are primary determinants of the environment in peri-infarct tissue and hence strongly influence the potential for neuronal plasticity. However, the specific roles of these cells and the timing of critical changes in their function are not well understood. Minocycline can protect against ischemic damage and promote recovery. These effects are usually attributed, at least partially, to the ability of this drug to suppress microglial activation. This study tested the ability of minocycline treatment early after stroke to modify reactive responses in microglia and astrocytes and improve recovery. METHODS: Stroke was induced by photothrombosis in the forelimb sensorimotor cortex of Sprague-Dawley rats. Minocycline was administered for 2 days after stroke induction and the effects on forelimb function assessed up to 28 days. The responses of peri-infarct Iba1-positive cells and astrocytes were evaluated using immunohistochemistry and Western blots. RESULTS: Initial characterization showed that the numbers of Iba1-positive microglia and macrophages decreased in peri-infarct tissue at 24 h then increased markedly over the next few days. Morphological changes characteristic of activation were readily apparent by 3 h and increased by 24 h. Minocycline treatment improved the rate of recovery of motor function as measured by a forelimb placing test but did not alter infarct volume. At 3 days, there were only minor effects on core features of peri-infarct microglial reactivity including the morphological changes and increased density of Iba1-positive cells. The treatment caused a decrease of 57% in the small subpopulation of cells that expressed CD68, a marker of phagocytosis. At 7 days, the expression of glial fibrillary acidic protein and vimentin was markedly increased by minocycline treatment, indicating enhanced reactive astrogliosis. CONCLUSIONS: Early post-stroke treatment with minocycline improved recovery but had little effect on key features of microglial activation. Both the decrease in CD68-positive cells and the increased activation of astrogliosis could influence neuronal plasticity and contribute to the improved recovery.

Rhesus brain gas tension and learned task performance responses to normoxic and hyperoxic breathing.

The impact of extended exposure to 21 and 100% O2 breathing mixtures on brain tissue oxygenation and performance of a learned task was studied in seven rhesus monkeys. Trained in the Sidman Avoidance Task, the animals were exposed to either breathing mixture for a period of 3.5 h in which brain tissue PO2, PCO2, and PN2 were observed. With 100% O2, PO2 and PCO2 rose significantly (less than 0.01 and less than 0.05, respectively) above control levels while PN2 fell significantly (less than 0.01) below its baseline value. With 100% O2 no statistically significant difference in gas tension values was shown for performance vs. nonperformance conditions. There was no difference in the number of shocks received with either breathing mixture. Lever press response frequency decreased throughout the experiment for both groups. During the last performance session, response frequency for the 100% O2 group leveled off, whereas for the 21% O2 group it continued to decrease.

Determination of cerebral and neck subcutaneous tissue PO2 with 3 and 4 +Gz in M. mulatta.

Four male and four female, fully conscious, M. mulatta were exposed twice each to diphasic accelerations of 3 or 4 +Gz (3 min) with a 10-min, 1-G intersession. One polarographic electrode was inserted intracortically and another one into neck subcutaneous tissue. With the first of two 3 or 4 +Gz exposures, cerebral PO2 fell from 15.11 to 5.73 and 2.92 torr, respectively; subcutaneous PO2 fell from 27.6 to 7.72 and 4.8 torr, respectively. Cerebral tissue exhibits post-G hyperoxia, related to initial stress intensity; even after the 10-min intersession, it is capable of mitigating the O2 desaturation effect of the second stress. Cerebral and subcutaneous tissue oxygen desaturation with G onset are comparable, but re-oxygenation in the latter tissue is apparently inertia-ridden and a simple relationship between the overall responses of cerebral and subcutaneous PO2 to +Gz acceleration could not be demonstrated.

Effects of increased ambient CO2 on brain tissue oxygenation and performance in the hypoxic rhesus.

Alterations of cerebral gas tensions and performance in response to hypoxia, with or without 5% CO2 in the ambient inspirate, were studied in eight conscious rhesus monkeys. The animals were trained to perform a lever press (Sidman) avoidance task. Physiological and performance data were obtained during exposures to normal (21% O2) and hypoxic (12, 10, and 8% O2--all with N2 balance) breathing atmospheres, with or without the addition of 5% CO2. With hypoxia, cerebral PO2 and PCO2 declined steadily, but a significant performance decrement was noted only at the 8% O2 level. With the addition of 5% CO2 to the inspired atmospheres, cerebral PO2 was elevated relatively but still declined as hypoxia intensified. Cerebral PCO2 and the avoidance task performance were sustained at near baseline values with the 5% CO2 inspirate.

A head enclosure for exposing monkeys to selected gas mixtures.

An acrylic head enclosure, permitting rapid exchanges of atmospheric gases, was developed for investigations of physiologic and performance responses of small nonhuman primates to graded hypoxic hypoxia and hypercapnia. The total system included a neck yoke plate which served both as a physical barrier to arm and hand movements in the head area and as a base on which to rest the head enclosure. The transparent, airtight, head enclosure permitted rapid exchanges of controlled gas mixtures and the execution of performance tasks in an apparently comfortable experimental environment.

Intracerebral oxygen and carbon dioxide tensions in the rhesus monkey.

Five rhesus monkeys (M. mulatta) were prepared by surgical placement of a stainless steel implant in the parietal area of the skull. The implant was designed to firmly hold a Teflon coated membrane of a medical mass spectrometer in an intracerebral position. Animals were consecutively exposed to varying ambient oxygen concentrations from 11% to 21%. Intracerebral PO2 and PCO2 and arterial PO2 were all stimultaneously recorded for computer analysis. Results indicate an almost linear relationship between cerebral PO2 and ambient PO2 as well as between cerebral PO2 and arterial PO2. With an ambient oxygen concentration of 21%, the cerebral Po2 and Pco2 were 13.5 +/- 3.5 mm Hg and 50.4 +/- 7.9 mm Hg, respectively. With an ambient oxygen concentration of 10.7%, these values decreased to 2.5 +/- 2.7 mm Hg and 38.0 +/- 5.6 mm Hg.

A chronic implant for intracerebral mass spectrometric gasometry in the rhesus monkey.

A cranial implant was developed to facilitate mass spectrometric gasometry in cerebral tissue of conscious primates. It is permanently fixed to the skull and allows repeated introduction of a Teflon diffusion membrane into the cerebrum. The implant is simple to manufacture and mount. It has been tolerated well with repeated introduction of the catheter for more than 6 mo in 10 rhesus monkeys. The preparation avoids the complication and artifacts of an acute surgical preparation for investigation of cerebral pO2 and CO2.