Biomedical subjects
Publications and source records attributed to H Acker.
A common capillary velocity of streaming blood, i.e. a common local flow combined with the geometry of the vascular elements already explains the measured left-shifted PO2 histogram. A slight variation of the vascular arrangement can also explain higher PO2 values without the assumption of a very high flow in the capillaries.
Changes of tissue pO2 levels in chronic subcutis wounds with and without heteroplastic tumour spheroid implants in thoraco-lumbal rat skinfolds covered by transparent chambers were followed in crosscut analysis by pO2 surface electrodes as a function of post-operative age using constant normoxic, hypoxic, post-hypoxic breathing conditions and intervals for each measured area. The tissue pO2 showed significantly higher values in the edematous phase of wound healing than in the granulation and differentiation phase. Tumour implantations induced even higher pO2 values in the edematic phase. This difference vanished in later phases. The hypoxic reactivity of non-tumour and tumour tissue improved with post-operative age indicating a sufficient vessel outgrowth.
Data of the intracellular electrolyte concentration of potassium and chloride in cultured muscle cells measured by x-ray analysis were compared by using the different activity coefficients with intracellular potassium and chloride activities measured with double-barrelled microelectrodes. By using an activity coefficient of 0.6, 95% of the potassium microelectrode measurements are in accordance with the x-ray analysis values, in spite of a scattering of the values. Membrane potential and intracellular potassium values are linearly related. x-ray analysis and ion-sensitive microelectrodes measured the cytoplasmic chloride in the same range. Taking into account known activity coefficients, an error of 25% must be assumed with the intracellular chloride measurements. However, x-ray analysis and ion-sensitive microelectrode investigations are reliable tools to study intracellular potassium and chloride changes, which play an important role in membrane characteristics.
Six different types of spheroids of both human and rodent origin were cultured, using the liquid-overlay technique. Oxygen gradients were measured with micro-electrodes, when the spheroids were attached to thin cover-glasses. The gradients were measured from the upper surface towards the center of the spheroids. Stable and reproducible gradients were obtained. Large variations were seen in the steepness of the gradients, depending both on the type and on the size of the spheroids. An interesting phenomenon was discovered. When some types of spheroids were cultured in a medium with a low oxygen pressure (medium equilibrated with gas containing 4-5% O2), the gradients continuously changed and became flatter. Detailed studies showed that most of the changes occurred within 2 days after the transfer to the low oxygen pressure. After 2 days, no further dramatic changes took place. This phenomenon was seen in two types of human glioma (U-118 MG and U-178 MG) and two types of embryonic, hamster lung-cell (V-79-379A and CHEL) spheroids. In the cases of human-osteosarcoma (U-393 OS) and human thyroid-cancer (HTh7) spheroids, no such changes could be seen. The low oxygen tension in the culture medium was chosen to mimic the environmental conditions in solid tumors.
To determine the importance of chemoreflex control for microcirculation, we investigated tissue pO2 and local flow on the surface of muscle, kidney, liver, and heart under systemic hypoxia and under isolated hypoxic perfusion of the carotid body in anaesthetized (pentobarbital) and artificially ventilated cats. Under systemic hypoxia, local blood flow mainly decreased in muscle and kidney, whereas liver and heart showed a flow distribution mainly with a flow decrease. Under systemic hypoxia, tissue pO2 decreased to values in the range of 0 to 10 Torr. A linear relationship was found between the pO2 change under hypoxia and the normoxic pO2 in the investigated organs, indicating local regulatory mechanisms of microcirculation. Isolated hypoxic perfusion of the carotid body caused a microflow increase in kidney in about 63%, in liver in about 55%, and in heart in all cases. Hypoxic perfusion of the carotid body caused pO2 increases in kidney and liver with mean pO2 values of 3 and 4 Torr, respectively, whereas, it produced no significant pO2 change in heart and muscle. The pO2 changes in kidney and liver significantly depended on the normoxic basic tissue pO2 level, indicating, in connection with local flow increases, a redistribution of local flow in these organs.
Type I cells of embryonic rabbit carotid bodies were cultured under normoxic (21% O2, 10% CO2, 69% N2) and hypoxic (5% O2, 10% CO2, 85% N2) conditions for two days. The mean membrane potential in the hypoxic cultivated type I cells (-27 mV) was significantly higher than in normoxic cultivated cells (-10 mV). The mean input resistance also had the tendency to increase under hypoxic conditions, from 19 m omega to 48 M omega. Results suggest that variations in membrane potential of type I cells due to oxygen pressure changes are an expression of the important role of these cells in oxygen sensing by the carotid body.
The adenosine triphosphate (ATP) level in the carotid body has often been discussed as the crucial step in the chemoreceptive process. Therefore, the ATP level of the cat carotid body was investigated with the aid of the bioluminescence method under different stimulation conditions. Under normoxic conditions an ATP level of about 0.087 nmol/glomus was measured, which is very low in comparison to other organs. The level did not change significantly, neither under hypoxic nor hypercapnic conditions. From these results we conclude that the primary effect of the chemoreceptive process in the carotid body cannot be explained by changes of the ATP level under different stimulation conditions.
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Different types of oxygen microelectrodes have been tested in measurements on cellular spheroids. The shape of the oxygen gradients varied strongly depending on size and type of the spheroids. No significant differences in the results were obtained when different types of electrodes were applied. All measurements were made in a perfusion chamber. The shape of the gradients did not vary with time in the perfusion chamber. The reproducibility was found good in repeated measurements using the same spheroid. No mechanical or chemical disturbances were seen during the penetration of the spheroids. Changes in the medium flow rate through the chamber did not drastically change the shape of the oxygen gradients. Almost no convection could be seen at the bottom of the chamber close to the spheroids. The composition of the medium was found to be of importance. Lock's solution containing glucose was found to be satisfactory. The potential signals in the double barrel electrodes allowed an accurate determination of the position when the electrode hit the spheroid surface. The information gained from microelectrode measurements in spheroids might be valuable for the understanding of effects of new tumor treatment modalities in which hypoxic cell sensitizers or high LET radiation are utilized.
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A new method was tested for studies of penetration of substances into tumorlike tissue. The penetration of the ions K+, Cl-, and Ca2+ through several layers of tumor cells was demonstrated by using double barrelled, ion sensitive microelectrodes with extra thin tip diameters. Spheroids consisting of human glioma, U-118 MG, and human thyroid cancer, HTh-7, cells were used as models of tumor tissue. A microelectrode was inserted into the center of a spheroid. Thereafter, the concentration of the test substance was increased in the surrounding medium. The change in concentration inside the spheroid was recorded and the penetration pattern evaluated. All three types of tested ions penetrated easily through the spheroids. The K+ ions penetrated most efficiently, and the Ca2+ ions showed the slowest penetration. The Ca2+ ions penetrated somewhat more slowly in the U-118 MG spheroids (which had rather small extracellular spaces) than in the HTh-7 spheroids (which had larger extracellular spaces). Ion sensitive electrodes, which are easily available, were used in this study only to demonstrate the principle. We hope that the method described can be used for penetration studies of various substances. For example, all substances that can be detected by enzyme microelectrodes could be studied. The main advantage of the method is that the complete penetration pattern can be studied as a function of time in individual spheroids. Previously described methods require histological procedures for each analyzed penetration time.
To determine the importance of dopamine and noradrenaline as neurotransmitters during chemoreception in the cat carotid body we investigated the contents of both compounds as well as the activity of dopamine-beta-hydroxylase (DBH) under different arterial PO2 and PCO2 conditions. The superior cervical ganglion was used as a control organ. In the carotid body and the ganglion an inverse relationship exists between the catecholamine content and the DBH activity. The carotid body has a high catecholamine content with a low DBH activity whereas the superior cervical ganglion has a low catecholamine content and high DBH activity. Hypercapnia did not produce any significant change in the catecholamine content or in the DBH content of the carotid body. However, in comparison with hyperoxia, hypoxia produced a significant change (p less than 0.05) in the noradrenaline content without changing the DBH activity. The dopamine content under these conditions did not change significantly. The results may indicate that the high catecholamine content of the carotid body is the result of a high retention and/or low rate of degradation rather than of a high rate of synthesis.
Human Glioma U-118 MG and hamster V-79-379A spheroids were used as models of the nodular structure often seen in poorly vascularized regions of solid tumours. K+, Ca2+ activities and membrane potentials were measured in the spheroids, using microelectrodes. Both stable and peak values were obtained. All values were included in the analysis to allow an accurate estimate of the relative number of electro-physiologically active cells at different depths in the spheroids. The relative number of active cells decreased drastically with depth although most cells in the corresponding regions looked morphologically viable. The amplitude values of potentials and K+ activities showed, however, no significant variations with depth. The Ca2+ activity varied largely between individual cells. The relative number of proliferative cells decreased with depth in the spheroids, in parallel to the decrease in the number of electro-physiologically active cells. The results indicate that the fraction of cells, in poorly vascularized regions, having normal metabolism might be fewer than what can be estimated only on a morphological basis. Thus, the amount of cells with the capacity to contribute to growth might be overestimated when inspecting histological sections.
A method has been described by which triple-barrelled ion-sensitive microelectrodes for simultaneous measurements of two extracellular ion activities could be produced with a tip diameter of 1-2 microns. The following combinations with two ion exchangers could be achieved: Ca++/K+, Ca++/Cl-, K+/Na+. The mean steepness for the potassium electrode was 55.8 mV, for the calcium electrode 26 mV, for the chloride electrode 48.8 mV, and for the sodium electrode 55.2 mV. The change of steepness varied 4%-5%/day and some of the electrodes could be used up to 5 days. The mean response time for all electrodes did not amount to more than 200 ms. An example is given for simultaneous in vivo measurement of both extracellular potassium and calcium activity.
1. The effects of electrical stimulation of the distal ends of cut preganglionic cervical sympathetic trunks and cut sinus nerves on carotid body total blood flow, local blood flow and tissue PO2 (Pt,O2) were studied in anaesthetized cats. 2. Sympathetic stimulation caused reductions, often marked, of the total blood flow through the carotid body, yet did not influence local flow or Pt,O2 recorded by electrodes whose tips lay in deep locations within the carotid body. Intraglomeral electrodes did respond to reductions of perfusion pressure and to perfusions of the carotid body with saline solutions. 3. Values of Pt,O2 recorded from superficial tissues of the carotid body were higher than those from deeper locations and were increased by blowing oxygen over the surface of the organ. In these locations, sympathetic stimulation increased Pt,O2. 4. An increase in the frequency of sinus nerve chemosensory discharges during sympathetic stimulation was obtained in the presence of an unchanged Pt,O2 recorded from deep glomeral locations. 5. Stimulation of the distal end of the sinus nerve increased the total blood flow through the carotid body but did not affect local flow or Pt,O2 recorded from deep glomeral locations. 6. It is concluded that autonomic nerves supplying the carotid body mainly influence arteriovenous anastomotic and/or other shunt vessels, with little control being exerted on vessels regulating the flow through the capillary network of the specific tissue. This investigation also indicates that the flow in the capillary network of the specific tissue is a small proportion of the total flow. Support is given to the view that autonomic nerves can influence chemoreceptor activity by non-vascular mechanisms.