Stephen C. Phillips

Natural loss of Purkinje cells during development and increased loss with alcohol

In untreated rats, degenerating Purkinje cells (P cells) were present from the 20th day of embryonic life (E20) to post-natal day 14 (PN14), but none was present on E17 or PN15. Ethanol increased the loss of P cells both before birth and up to PN6 or 8, but from PN10 no increased loss was seen. Most degenerating P cells were located below the P cell line. On PN3 ethanol killed some P cells in 12 h, and the first ultrastructural change was a dilatation of the nuclear double membrane after 8 h. Among the surviving P cells, natural cell death continued. Pentothal anesthesia on PN3 did not damage P cells. Exposure to ethanol from E12 to PN5 resulted in a large loss of P cells and retarded the foliation of the cerebellum.

Is the inferior colliculus and obligatory relay in the cat auditory system

The status of the inferior colliculus of the cat as an obligatory relay in the ascending auditory pathway was examined by attempting to infiltrate totally the fibres of the brachium of the inferior colliculus on one side with horseradish peroxidase. Following a transport time of 24 h, alternate sections from thalamus to caudal brainstem were reacted with a sensitive histochemical method to reveal tracer reaction product. Results for three cats revealed that the inferior colliculus is an obligatory relay for the overwhelming majority of axons comprising the lateral lemniscus and originating in the cochlear nucleus and superior olive.

Chronic consumption of alcohol by adult mice: effect on hippocampal cells and synapses.

For 4 months C57 black mice were fed a nutritionally complete diet containing 9% alcohol or isocaloric sucrose and killed then or after 4 months recovery on standard food pellets. The number of cells in a thin plastic section of hippocampus was unchanged in field CA1 by alcohol exposure but was reduced 9% during withdrawal from alcohol. Electron microscopy was used to count synapses among the basal dendrites and no significant change was found in any treatment group. The spine heads were measured and found to be smaller in the alcohol group than in the sucrose group; many of the spines (in the alcohol group) were too small to be visible with the light microscope.

Age-dependent susceptibility of rat cerebellar Purkinje cells to ethanol exposure☆

It is known that a proportion of cerebellar Purkinje cells do not complete development in the normal rat. In this study neonatal rat pups were treated at various stages of Purkinje cell development with ethanol vapour. We observed an increased rate of Purkinje cell loss at postnatal day 3, yet identically treated littermates had a normal complement of Purkinje cells compared to age-matched controls at 47 days of age. Single day ethanol exposures during Purkinje cell ontogenesis seems to accelerate a natural loss of Purkinje cells without a permanent loss persisting to adult life.

Does ethanol damage the blood-brain barrier?

The permeability of the blood--brain barrier has been measured using a technique which is independent of blood flow and is sufficiently accurate to monitor the penetration of weakly permeant substances. The permeability of the blood--brain barrier to [14C]sucrose has been measured in rats anaesthetised with either urethane or pentobarbitone (Nembutal). The values obtained from urethane-anaesthetised untreated rats were slightly lower, thus demonstrating the suitability of urethane as an anaesthetic for blood--brain barrier experiments. The permeability of the barrier has been measured in rats which had been drinking 7.5% ethanol for 6 months, or had been administered an anaesthetic dose of ethanol, or both. No statistically significant difference was found between the permeability measurements in rats subjected to any of these treatments. Positive controls in which 0.3 ml of a 30% ethanol solution was injected into the internal carotid artery demonstrated the sensitivity of the employed technique. Thus it was found that the blood--brain barrier does not weaken with respect to sucrose when the blood ethanol concentration reaches an anaesthetic level.

Qualitative and quantitative changes of mouse cerebellar synapses after chronic alcohol consumption and withdrawal

For 4 months C57 mice were fed a nutritionally complete diet containing 9% alcohol or isocaloric sucrose and killed then or after 4 months recovery on standard food pellets. Electron microscopy was used to count synapses among the cerebellar Purkinje cell dendrites and a significant 13% reduction was found in the alcohol recovery group. The shape of the postsynaptic density material was studied and a significant 11% increase in length found in the alcohol group. The smaller thickness of the postsynaptic material in animals of the alcohol recovery group may be representative of new synaptic formation.

Structural plasticity of developing optic synapses under different lighting conditions

Four-month-old male hooded rats were reared from birth under constant light and darkness conditions. Changes in the amount of postsynaptic density material in the optic synapses in the suprachiasmatic nucleus of these rats were compared with animals maintained under routine light-dark (12 h) cycles. The thickness of postsynaptic density material was found to be significantly greater in dark-reared rats relative to light-reared animals. The plasticity of this structure may have functional implications in the sensitivity of postsynaptic response. There was no significant difference in the lengths of the synaptic apposition and the size of boutons.

The short-term toxicity of ethanol to neurons in rat cerebral cortex tested by topical application in vivo, and a note on a problem in estimating ethanol concentrations in tissue

In rats anesthetized with ethanol 4.0 g/kg i.p. the dura overlying the parietal cortex was exposed and superfused with 100% ethanol for 1 h. After 6 days survival the underlying cortex was stained with a silver method that is selective for degenerating axons and their terminals. No degeneration was found in the superfused cortex, although heat-lesioned tissue stained concurrently showed axonal degeneration and so validated the technique. Electron microscopy after 3-20 days survival did not show any degeneration, and synapses of normal appearance were present immediately beneath the cortical surface. In other rats the ethanol concentration in the superfused tissue was assayed in 0.4 mm thick discs sectioned with a vibratome from a 4-mm diameter core cut with a trocar from the cortex immediately after 1 h of superfusion. The ethanol was eluted in 2% TCA, and an aliquot assayed enzymatically. A second elution of the tissue disc contributed a further 5% of the ethanol content indicating a partition coefficient for ethanol between wet brain tissue and 2% TCA of about 10. The total concentration of ethanol in the superficial cortex was found to be about 0.82 M or 3.8%. This estimation was confirmed by superfusion with 14C-labelled ethanol and scintillation counting. Thus neurons in the cerebral cortex did not degenerate after exposure for 1 h to a concentration of ethanol that was 3 times greater than the concentration that causes death in a rat by paralysis of the respiratory centre (1.2%).

Natural variation in the blood-brain barrier

Measurement of the product of the permeability × vascular surface area (PA) of the blood-brain barrier to [14C]sucrose has shown a significant sex difference in an inbred strain of rats. Moreover, male rats of the Sprague-Dawley strain differ significantly from male DA Agouti rats, but not from male guinea-pigs. Thus the varying estimates of PA in the recent literature may be due to genetic variation rather than to differences of experimental techniques.

Cytoprotective value of lysine, penicillamine, and pyridoxal phosphate against the neurotoxicity of acetaldehyde

Rats which received acetaldehyde by intraperitoneal injection on a single occasion sustained neural degeneration in the cerebral cortex detectable with both light and electron microscopy. The degeneration was more intense and included the hippocampus when acetaldehyde exposure was given on 5 consecutive days in the form of ethanol vapor inhalation and disulfiram injections. Pretreatment of animals with a combination 2.5 g lysine/kg plus 50 mg pyridoxyl phosphate/kg did not alter the degree of degeneration response to either a single injection of acetaldehyde or 5 days acetaldehyde treatment. Penicillamine injections (1.2 g/kg) did offer some (but not complete) cytoprotective value against the neurotoxicity of acetaldehyde. This cytoprotective action was effective at concentrations of acetaldehyde which are not distant from clinically observed concentrations.