Joseph DiMattio

Effects of changes in serum osmolarity on bulk flow of fluid into cerebral ventricles and on brain water content

SummaryThe effects of changes in serum osmolarity on the rate and osmolarity of bulk flow of fluid into the cerebral ventricles and on cortical white and grey matter water content were studied in cats. Bulk flow rates and osmolarities were measured during ventriculocisternal perfusion both before and after intravenous infusion of glucose solutions. Infusions of glucose in concentrations greater than 6% decreased fluid bulk flow rate and its osmolarity. Glucose in concentrations less than 6% increased fluid bulk flow rate and decreased its osmolarity. Bulk flow rate and serum osmolarity were found to be linearly related with a coefficient of osmotic flow of −0.835 μl/min per mOsm/l. At the extremes of induced serum osmolarities, (290 and 360 mOsm/l) bulk flow rate was either increased by 120% or completely inhibited. Effluent osmolarity also increased proportionately to serum osmolarity (0.338 mOsm/l per mOsm/l). When compared to controls, cortical grey and white matter water content increased by 1.9% and 2.9%, respectively, when the infused glucose concentration was 2.5% or less, and decreased by 1.8% and 2.9% when the concentration was 10% or more. The results of these experiments suggest that the increased bulk flow comes from the brain, rather then directly from the blood.

A model for transepithelial ion transport across the isolated retinal pigment epithelium of the frog

The contribution of chloride ion movement and sodium and bicarbonate concentrations to the net current across the isolated choroid-retinal pigment epithelium (RPE) of the bullfrog were studied. The presence of a ouabain-sensitive Na+/K+-pump on the retinal side was confirmed and complete inhibition of this pump with Na+ removal and ouabain treatment abolished nearly all the RPE transepithelial transport and SCC suggesting that all ionic transport was dependent on sodium. It was found that apical to basal (AB) chloride flux accounted for 26 +/- 2% (mean +/- S.E.M.) of the short circuit (SCC). Results suggest that AB bicarbonate and/or basal to apical (BA) hydrogen ion net transport accounts for 38 +/- 2% of the SCC while BA sodium is presumably responsible for the remaining 34% of the SCC. Transport was inhibited by apical administration of known chloride inhibitors. Trans-RPE 36Cl flux measurements indicate that furosemide (10(-4) M) and SITS (10(-3) ) decrease the retinal-choroid flux. Results suggest that net transport of chloride and bicarbonate are independent of each other and additive. It was found that a bicarbonate-free preparation was relatively unaffected by changes in pH (5.5-8.5) indicating that pH has little, if any, effect on sodium or chloride current in this range. A model is presented which is compatible with the various data. It is suggested that along with the apical Na+/K+-ATPase pump, there exists an apical Na+/Cl- -co-transport system which is driven by the established sodium gradient. Moreover, this pump established sodium gradient is postulated to drive a Na+/HCO3- -co-transport system tentatively placed on the retinal side of the RPE.

Glucose transport into the ocular compartments of the rat

An exchange model was developed for the study of the transfer between a central plasma compartment and the aqueous and vitreous compartments in the rat eye. The solutes investigated include sucrose, urea, 3-O-methyl- d -glucose, d -glucose and l -glucose. For the substances studied, rate parameters were used which enabled ocular concentration functions to be evaluated from various plasma functions. Rate transfer constants were evaluated from plasma functions and compartments significantly faster than does urea. d -glucose is transported six times faster than l -glucose, indicating that the transport mechanism is stereospecific. The saturability of the d -glucose transport system was explored by elevating the plasma concentration of unlabeled d -glucose. The system was found to be saturable. A Michaelis-Menten analysis of transport vs. Plasma concentration revealed the Km to be 6·08 m m and 4·64 m m and the Vm to be 0·0225 and 0·0238 mmoles/min (for the aqueous and vitreous repectively). Relative high glucose concentrations (50 m m ) were required for saturation. We conclude that transport of glucose into the ocular fluids is consistent with a carrier-mediated facilitated diffusion mechanism which is stereospecific and saturable.

The effects of serum osmolarity on cerebrospinal fluid volume flow

Abstract The effects of changes in serum osmolarity on cerebrospinal fluid (CSF) formation were studied in cats. CSF production rates were measured by ventriculocisternal perfusion both before and after intravenous infusion of glucose solutions. Infusion of glucose, hyperosmolar with respect to serum, increased serum osmolarity and caused a decrease in CSF formation rate; conversely, infusion of hypoosmolar solutions lowered serum osmolarity and increased CSF formation. CSF production and serum osmolarity were found to be linearly related. A 1% serum osmolarity change resulted in a 6.7% change in CSF formation. CSF formation increased by 130% with a serum osmolarity of 265 m0sm/1 and was undetectable with serum of 380 m0sm/1.

Facilitated glucose transport across the retinal pigment epithelium of the bullfrog (Rana catesbiana)

Transport studies of glucose analogs [3H] 3-O-methyl-D-glucose (mD-glu) and L-[14C]glucose (L-glu) across the isolated retinal pigment epithelium (RPE) of the bullfrog was undertaken to determine whether the glucose transport mechanism was dependent upon the postulated ion-transport scheme and/or whether glucose transport is insulin-mediated. In addition, metabolic inhibitors were tested to explore the energy requirements of glucose transport across the RPE. Flux studies of mD-glu and L-glu performed with mounted RPE tissues, with short circuit current (SCC) and potential difference (PD) monitored via automatic voltage clamp apparatus, indicate that transport is clearly stereospecific with D-glucose being transported at least 13 times faster than L-glucose. The system was found to be saturable with a Km of about 24 mM glucose and Vmax of 1400 nmol cm-2 hr-1. Unidirectional Michaelis-Menten constants indicate that the RPE glucose carrier is accessible for transport from either the choroid or retinal side and a bidirectional facilitated diffusion mechanism is suggested. Insulin had no effect on either ion transport (SCC) or glucose transport (passive or facilitated). Both aerobic and anaerobic energy inhibitors decreased ion transport to less than 25% of control, but had little effect, if any, on glucose transport across the isolated RPE. Sodium iodoacetate decreased ion transport by 90% of control, but a much slower decrease in facilitated glucose transport of 22% of control suggests that carrier energy requirements, if any, are not direct or immediate. Osmotic studies performed with sucrose and glucose suggest that elevations in osmolarity increase passive glucose movement and decrease facilitated glucose-transport rates. Glucose was found to be much more detrimental to glucose transport than sucrose, suggesting that at high concentrations molecular glucose decreases facilitated transport and increases passive glucose movement by a mechanism other than can be accounted for by osmotic considerations. A model for RPE glucose transport, consistent with current data, is proposed which translocates D-glucose, via an alternating conformational change of the glucose carrier. This carrier does not require a direct supply of metabolic energy, nor a functioning ion-transport mechanism. At a given moment, a single binding site for D-glucose is postulated to be available on either side of the RPE membrane for glucose translocation, although binding site affinity for glucose could differ on each side.

Glucose Transport Across Ocular Barriers of the Streptozotocin-Diabetic Rat

The transport kinetics across the plasma-aqueous and plasma-vitreous barriers were studied in normal and long-term streptozotocin-diabetic rats, using trace amounts of [14C]-L-glucose and [3H]-3-O-methyl-D-glucose. The former is passively transported while the latter uses the same transport-facilitating system as D-glucose. Transport rates of L-glucose were significantly higher in the diabetic rats, with ocular entry rates from the plasma being increased by 69% across both barriers. Thus, the data indicate that in experimental diabetes the passive permeability of the bloodocular barriers is significantly increased. By contrast, calculated transport rate constants for 3-O-methyl-O-glucose, when adjusted for the hyperglycemia and the increased passive glucose movement, are not altered in the diabetic animal. Nevertheless, there is actually more mass D-glucose movement due to the prevailing hyperglycemia. The present study suggests that although streptozotocin diabetes alters plasma-ocular glucose transport, there is no direct impairment of glucose carrier function. Alterations in transport occurred at both ocular barriers, suggesting that involvement is general and that both the retinal pigment epithelium and the ciliary epithelium may be affected by the diabetes. It is unknown whether the increase in passive movement is related to the prevailing hyperglycemia or to insulin deficiency or other unknown factors.

Reduced ocular glucose transport and increased non-electrolyte permeability in rats with retinal degeneration (RCS)

The transport kinetics across the plasma-ocular barriers of labeled molecules including urea, D-glucose, 3-0-methyl-D-glucose, L-glucose, and sucrose were studied in adult RCS rats and compared to control albino Sprague-Dawleys (SD). Results indicate that substances that passively cross the plasma-aqueous and plasma-vitreous barriers (urea, sucrose, L-glucose) do so more readily in the RCS rat especially via the trans-retinal route. By contrast, D-glucose, which penetrates the ocular barriers of the control rat by a carrier-mediated mechanism was found to cross the ocular barriers of the RCS rat at a reduced rate. Thus, the ocular barriers of the RCS rat with well developed retinal degeneration demonstrate an increase in passive permeability and a reduction in ability to perform their membrane transport function (D-glucose). The permeability defect was more pronounced in the blood-vitreous barrier than in the blood-aqueous barrier.

Cerebral fluid dynamics and brain regional blood flow in experimental hydrocephalus.

Cerebral blood flow was measured by the indicator fractionation technique in normal, acute hydrocephalic, chronic compensated hydrocephalic and craniectomized hydrocephalic cats. In the five normal cats the mean total brain blood flow was 136.1 ml/min/100 g dry weight. The six acute hydrocephalic animals demonstrated a relatively uniform 22% reduction in total blood flow. In eight chronic hydrocephalic cats CBF increased to the point where there was only an overall 7% decrease. In three hydrocephalic and craniectomized cats the CBF was reduced by 30.6%. In the acute phases there was a decrease in the number of blood vessels. Chronic compensated hydrocephalic brains had somewhat more vessels than the normal, whereas the craniectomized, massively hydrocephalic brain had a dramatic increase in both the number and caliber of blood vessels. These results clearly demonstrate that in acute obstructive hydrocephalus in cats, there is a significant decrease in CBF. The blood vessels revert to normal in shunted cats.

Preparation and Analysis of 4-lodoantipyrine-(I 131 )

4-Iodoantipyrine was prepared from antipyrine according to the reaction equation: 3C11H12N2O+2KI+KIO3+3HCl→3C11H11N2OI+3KCl+3H2O The product was separated from the reaction mixture and purified by recrystallization. The iodoantipyrine was labeled with I131 by means of an exchange reaction with NaI131. It was separated from the unbound radioactivity by anion exchange chromatography. The purity of the product was determined by cellulose thin layer chromatography using ethanol-chloroform-water (45:45:10) as the solvent system. The purity of the 4-iodoantipyrine-I131 decreased with time due to a splitting off of the I131 atom and subsequent formation of NaI131. This spontaneous deiodination was best minimized by storage in methanol at low temperatures.