Carole H. Latker

Altered Blood-Nerve Barrier Permeability to Small Molecules in Experimental Diabetes Mellitus

Permeability-surface area products (PA) were determined with a quantitative in vivo injection technique at the blood-nerve barrier of tibial nerve, and at the blood-brain barrier, in control and streptozotocin-induced diabetic rats. The PA product for [14C]mannitol at the blood-nerve barrier was increased by 100% in diabetic animals, 3.12 ± 0.15 ± 10-5 mls-1·g-1, compared with controls, 1.61 ± 0.10 ± 10-5 ml·s-1·g-1. In contrast, PA for [14C]mannitol at the blood-brain barrier was unaltered in the diabetic animals. Following intravenous injection, no leakage of microperoxidase across the perineurium or endoneurial vessels of diabetic rats could be demonstrated by morphological techniques. Nerve blood-space, as determined with intravenous [3H]inulin, and blood-nerve barrier surface area as determined by morphometric methods, did not differ in diabetic when compared to control animals. Thus, the calculated permeability coefficient for [14C]mannitol at the blood-nerve barrier was about 100% greater in diabetic nerve compared to control nerves. The increased permeability was accompanied by a 7% increase in nerve-water content and a 32% decrease in motor-nerve conduction velocity. The results demonstrate a specific vulnerability of nerve as compared to brain in an animal model of diabetes mellitus. Chronically altered permeability to small water-soluble molecules reduces the protective effect of salt impermeability at the blood-nerve barrier against nerve edema, and may be an important pathogenic mechanism in diabetic neuropathy.

Cerebral metabolism of plasma [14C]palmitate in awake, adult rat: Subcellular localization

Following intravenous injection of [U-14C]palmitate in awake adult rats, whole brain radioactivity reached a broad maximum between 15–60 min, then declined rapidly to reach a relatively stable level between 4 hr and 20 hr. At 44 hr total radioactivity was 57% of the 4 hr value (p<0.05). About 50% of palmitate which entered the brain from the blood was oxidized rapidly, producing14C-labeled water-soluble components which later left the cytosol. Radioactivity in the cytosolic fraction peaked at 45 min and then declined, coincident with the decline in total brain radioactivity. Membrane fractions were rapidly labeled to levels which remained relatively stable from 1 to 44 hr. Increases in the relative distributions of radioactivity were seen between 1 and 4 hr for the microsomal and mitochondrial fractions, and beyond 4 hr for the synaptic and myelin membrane fractions (p<0.05). Radioactivity in membrane fractions was 80–90% lipid, 5–13% water-soluble components and 3–17% protein. The proportion of label in membrane-associated protein increased with time. Proportions of radioactivity in the combined membrane fractions increased from 65% to 76% to 80% at 4, 20 and 44 hr, respectively. The results show that plasma-derived palmitate enters oxidative and synthetic pathways to an equal extent, immediately after entry into the brain. At and after 4 hr, the radiolabel resides predominantly in stable membrane lipids and protein. Brain radioactivity at 4 hr can be used therefore, to examine incorporation of palmitate into lipids in vivo, in different experimental conditions.

Assessment of the permeability of the blood-retinal barrier in hypertensive rats.

We assessed the permeability surface area products at the blood-retinal barrier and blood-brain barrier to sucrose (molecular weight, 340) and microperoxidase (molecular weight, 2000) following acute hypertension induced by metaraminol in Wistar-Kyoto rats (controls) and during chronic hypertension in spontaneously hypertensive rats. In acute hypertension, the permeability surface area product for sucrose was increased at the blood-retinal barrier and at the blood-brain barrier over control values (p less than 0.02), and the vessels became leaky to microperoxidase. In chronic hypertension, the permeability of the blood-retinal barrier to sucrose was increased over that in control animals (p less than 0.02), whereas the permeability of the blood-brain barrier was unaffected. Neither barrier leaked microperoxidase. These results indicate that the blood-brain barrier and the blood-retinal barrier are similarly affected in acute hypertension and that in chronic hypertension, the blood-brain barrier is unaffected whereas the blood-retinal barrier is rendered more permeable to small, but not large, solutes.

Vesicular profiles in frog perineurial cells preserved by rapid-freezing and freeze-substitution

Vesicular profiles in perineurial cells of frog peripheral nerves were examined in tissues preserved by rapid-freezing or by conventional chemical fixation. Tissues were processed immediately after removal from the animal or after remaining in Ringers for several hours. Vesicular profiles were present in perineurial cells in all experimental groups, demonstrating that they are not an artifact of chemical fixation. However, variations in their morphology correlated with the different preservation techniques.

Perineurial permeability and endoneurial edema during Wallerian degeneration of the frog peripheral nerve

Perineurial permeabilities to [3H]sucrose and [14C]dextran (MW = 70,000), and water content, conduction velocity (CV) and maximum amplitude (MAP) of the compound action potential, were determined in Wallerian degenerated nerves (sciatic or tibial) of the frog and compared with values in the contralateral uncut nerves. Three days after transection of the lumbosacral plexuses, about 2 cm proximal to the sciatic nerve, mean water content of the sciatic nerve was significantly higher than in the contralateral uncut nerve. After 10 days, the degenerating sciatic nerve showed significant increases in the mean perineurial permeabilities to [3H]sucrose and [14C]dextran when compared to values in the contralateral nerve. Means MAPs and CVs were significantly decreased. At 21 days and after, no compound action potential was detected and perineurial permeability and nerve water content had increased further. Decreases in mean MAPs and CVs and permeability increases of the perineurium were less in degenerating tibial nerves than in degenerating sciatic nerves. It is concluded that following transection, (1) Wallerian degeneration produces an irreversible increase in perineurial permeability, (2) the increase of perineurial permeability follows a proximodistal gradient, and (3) the frog peripheral nerve develops endoneurial edema during Wallerian degeneration as do degenerated nerves of mammals.

The morphology of pial blood vessels of the frog, preserved by rapid freezing and freeze substitution.

Vesicular profiles in endothelial cells of frog meninges were examined in tissues preserved by rapid freezing or conventional chemical fixation. Tissues were frozen immediately after removal from the animal or after remaining in Ringers for several hours. Vesicular profiles with an average diameter of 240 nm were present in the endothelial cells in all experimental groups, demonstrating that they are not an artifact of chemical fixation or incubation in vitro. However, their concentration and morphology varied with the different preservation techniques.

Adrenergic innervation of blood vessels in rat tibial nerve during Wallerian degeneration.

SummaryAdrenergic innervation of blood vessels in rat tibial nerve during Wallerian degeneration was examined, using the formaldehyde-induced histo-fluorescence method. The left sciatic nerve was transected at the level of the sciatic notch, whereas the right sciatic nerve was left intact and used as control. At 1, 3, 7, 14, 42, 56 or 84 days after transection, the tibial nerves of the transected and contralateral sides were exposed. Pieces of each nerve were used for light microscopy or for examination of adrenergic innervation with the fluorescence microscope. One day after transection, no adrenergic nerve fiber was observed in the endoneurium of the transected nerve. After 3 days, adrenergic innervation of small-and medium-sized arterioles in the epi-perineurium was absent, and after 7 days no fibers were visible around large arterioles. Fluorescent fibers were not detected even at 84 days post-surgery. It is concluded that adrenergic innervation of blood vessels in the rat tibial nerve is irreversibly lost after permanent axotomy, and that adrenergic regulation of nerve blood flow may also be lost.