G. Duncan

Ion analyses of human cataractous lenses

Abstract The sodium, potassium, calcium, magnesium and chloride concentrations have been determined in 52 human cataractous lenses. Three main types of association between concentrations and cataract type were found. Group A, mainly nuclear cataracts, had a near normal range of concentrations. Group B, a mixture of cataract types, including mixed cataracts, had a significantly raised sodium and lowered potassium concentration, but particularly a raised calcium concentration. Group C, mainly cortical and mature cataracts, had grossly increased sodium and calcium concentrations, an increased chloride concentration and a very low potassium level. In all groups the magnesium concentration remained relatively constant. The role of calcium is particularly interesting. In pure nuclear cataracts its concentration is relatively normal and below the equilibrium concentration, but when there is cortical involvement very high calcium levels are found. The values, which are above the calculated equilibrium concentration, indicate that binding of calcium occurs. Group B and C cataractous lenses also showed a significant decrease in dry weight.

REGULATION OF CALCIUM FLUXES AND THEIR REGULATORY ROLES IN PANCREATIC ISLETS

It is today considered as crystal clear that calcium plays an essential role in the regulation of insulin release by the pancreatic B-cell. Some of the major issues concerning such a role are as follows: ( i ) what is the detailed mechanism by which secretagogues are susceptible to influence the handling of calcium in the B-cell; (ii) what is the nature and location of the critical pool of calcium that controls insulin release; (iii) what is the relative and respective contribution of calcium influx, efflux, and subcellar distribution in the regulation of such a pool; and (iv) how does calcium influence the process by which secretory granules migrate to the cell boundary and are extruded via exocytosis in the interstitial fluid.’ In the present report, for the sake of clarity, we will restrict the discussion of these questions to the process of glucose-induced insulin release, with the main emphasis on the possible significance of passive ionophoretic movements. The process by which glucose provokes insulin release can be viewed as a sequence of three major events, namely: ( i ) the recognition or identification of glucose by the B-cell; (ii) the subsequent remodelling of cationic fluxes; and (iii) the activation by calcium of an effector system controlling the migration and exocytosis of secretory granules.2 The role of calcium in insulin release is here considered within the framework of such a sequential view.

The site of the ion restricting membranes in the toad lens.

Two experimental approaches have been used to determine whether the internal fibres of the amphibian lens have intact membranes. One approach involves an analysis of the potential transients obtained when potassium is substituted for sodium in the external medium and the other involves a correlation of lens electrical resistance with depth of penetration of the measuring electrodes. The half-times of the potential transients were in the region 100–120 sec and taking 4 × 10−6 cm2 sec−1 as the diffusion coefficient of potassium, these values correspond to a diffusion path length of 200–350 μ. As an “unstirred layer” of the surrounding medium probably extends for 100–150 μ from the lens surface, the potential-determining membranes lie at most 100–200 μ below the surface. As the diffusion of potassium through the capsule and through the extracellular space of the lens is probably much slower than in free solution, this distance may well be reduced to 10–20 μ. The resistance, measured by the two-electrode method, was found to be independent of the depth of the electrodes below the surface. There was also no marked difference in the resistances of the anterior and posterior faces of the lens. These resistance results also suggest that the ion-limiting membranes lie near the surface.

A comparison of ion concentrations, potentials and conductances of amphibian, bovine and cephalopod lenses

1. The concentrations of sodium, potassium and chloride in frog and bovine lenses showed a normal intracellular ion distribution with the sum of the internal cations approximately equal to the external sum. In the cephalopod lens, however, the sum inside was much lower than that outside.

Classification of human senile cataracts by nuclear colour and sodium content

Abstract Human cataractous lenses were photographed in vivo prior to surgery and then in vitro after extraction. They were also analysed for sodium concentration and protein content. The lenses were then classified according to a dual system taking into account the extent of nuclear involvement (colour groups I to V) and electrolyte changes (sodium bands A to E) in each lens. The protein data from lenses classified according to the above dual scheme confirmed that the colour of the lens is closely associated with the water-insoluble protein content of the lens but further demonstrated that an increase in the sodium concentration (osmotic involvement) is associated with a dramatic fall in the water-soluble and total protein content. As the osmotic and nuclear mechanisms lead to quite different changes in protein distribution and as most cataracts are mixed in nature, a dual classification system is essential in any study where it is intended to discriminate between the two mechanisms. It was found that both the in vivo and in vitro photographic methods gave a consistent measure of nuclear involvement. However, it was not always possible to relate the changes in lens sodium to the extent of light scattering seen either in the in vivo or in vitro photographs.

Effect of glucose on K+ handling by pancreatic islets

Data are presented which confirm and extend conclusions derived from previous experiments carried out with 86Rb+. They suggest that glucose, by reducing the permeability of the plasma membrane to effluent K+, increases the concentration of K+ in the islet cells and under steady-state conditions, reduces proportionally the fractional turnover rate of the cellular K+ pool. Glucose, however, does not increase the inflow-outflow rate of K+ into the islet cells. The mechanism by which glucose affects the permeability of the plasma membrane to effluent K+ remains to be elucidated.

Histamine and ATP mobilize calcium by activation of H1 and P2u receptors in human lens epithelial cells.

1. Cytoplasmic calcium concentration ([Ca2+]1) of single superfused tissue‐cultured human lens epithelial cells (HLEC) was monitored using the fluorescent dye fura‐2; the resting values were low and stable for several hours ([Ca2+]i = 96 +/‐ 20 nM; mean +/‐ S.D., n = 16). 2. Continuous superfusion with either ATP or histamine (0.1‐10 microM) produced regular oscillations in [Ca2+]i that could be maintained for a short time in the absence of external calcium. 3. Short (30 s) pulses of histamine (0.1‐100 microM) induced a transient rise in [Ca2+]i, the time course of which was insensitive to the removal of external calcium. The rate of rise and the amplitude of the response were very sensitive to agonist concentration, whereas the rate of recovery was relatively constant. 4. The responses to long pulses of histamine (> 100 s) consisted of an initial transient followed by a maintained [Ca2+]i which returned to baseline on removal of external calcium. 5. The kinetics of the responses to short and long pulses of ATP (0.1‐100 microM) were very similar to those of histamine and showed a similar sensitivity to the presence or absence of external calcium. 6. The histamine responses were abolished by triprolidine (1 microM), but unaffected by ranitidine (1 microM), indicating that an Hi receptor subtype is activated by histamine. 7. The ATP responses were reversibly inhibited by suramin and the potency sequence for a range of agonists was ATP = UTP = ATP gamma S > ADP = GTP >> AMP = adenosine, indicating that activation of a P2u receptor subtype was responsible for the increase in [Ca2+]i. 8. Both histamine and ATP responses were abolished by thapsigargin (100 nM), confirming that calcium release from intracellular stores was responsible for the initial peak of the response. Application of either agonist during the plateau phase of the thapsigargin response often led to a marked, but reversible, decline in [Ca2+]i, indicating the presence of a further, normally hidden, calcium regulatory factor associated with the presence of the agonist. 9. Maximal concentrations of either histamine or ATP totally emptied the calcium store as a subsequent application of the other agonist (or thapsigargin), in the absence of external calcium, failed to induce a further increase in the calcium signal.

Calcium-induced opacification and loss of protein in the organ-cultured bovine lens

A long-term system of organ culture for bovine lenses was used to investigate the effect of osmotic stress on lens opacification and crystallin loss. Lenses were pre-incubated in control medium containing L-[U-14C]tyrosine so that labelled crystallins were produced. The fate of these crystallins was studied in relation to two forms of osmotic stress. The addition of either ouabain or EGTA to the medium induced severe osmotic swelling and disturbance of the lens monovalent cation balance, but only the former treatment was followed by an increase in lens calcium. The changes due to osmotic stress were accompanied by loss of transparency and protein only in the lenses with increased calcium. Both opacification and increased calcium were found largely to be confined to the outer cortical fibres. Protein loss increased with time as lens calcium continued to increase. The protein recovered from the incubation medium was characterized by gel filtration and immunological techniques. The first protein detected was beta L-crystallin, and this formed the major part of the lost protein throughout, although alpha- and gamma-crystallins were detected at a later stage. Increased calcium also resulted in a change in the susceptibility of the crystallins to aggregation, since there was an increase in [14C]tyrosine incorporated into the lens high-molecular-weight (HM) fraction after exposure to ouabain, but not after exposure to EGTA. The relevance of these findings to human cataract is discussed.

The aging human lens: structure, growth, and physiological behaviour

The aging human lens has been the subject of intense research over the past 20 years, for a number of quite disparate reasons. The fact that the incidence of cataract rises exponentially with age after 50 years1 provides the driving influence for much of the effort, but the unique accessibility, homogeneity, and basic simplicity of structure of the organ itself makes it a fruitful system for fundamental studies of tissue growth, development, and differentiation.2-4 Images of the whole human lens in vivo have been available for detailed analysis since the introduction of the quantitative slit lamp (or Scheimpflug) camera (Fig 1). This has been invaluable in providing an understanding of the changes in shape and optical properties both of the ‘normal’ and cataractous aging lens.5-7 The lens is also accessible as an in vitro system of study through the provision of donor eyes for corneal transplant and general research. Since the lens has no direct blood supply, it survives well both in the globe itself and in organ culture media.8-10 Whole cataractous lenses were also once routinely available before the advent of extracapsular cataract extraction (ECCE) with intraocular lens implantation and in the past there have been combined slit lamp and in vitro studies which have correlated changes in light scatter and absorbance with specific alterations in ion and protein levels (Fig 1 and Marcantonio et al 11 and Hockwin et al 12). Figure 1 Images of normal human lenses (A, B, and C), posterior polar cataract (D, E, F), and pure nuclear cataract (G, H, I). Note that slit lamp camera images (A, D, G) all have a scattering reflect artefact (small white rectangle). The normal subject (A) was 40 years of age and the accompanying in vitro grid photographs (B) and polarising images (C) were …

Aqueous humour glucose concentration in cataract patients and its effect on the lens

The glucose concentrations of the blood plasma and aqueous humour were measured in 56 cataract patients. The sodium concentration and colour of the lenses were determined after extraction. The mean plasma and aqueous glucose levels were 5.8 and 3.2 mM respectively in non-diabetic patients, while the values for diabetics were 14.2 and 7.8 mM. The sodium concentration of the lenses from non-diabetic patients appeared to consist of two distributions around 30 and 170 mM, corresponding to nuclear and cortical cataracts respectively. Only two lenses from the non-diabetic patients had sodium concentrations in the range 60-120 mM. In diabetic patients, however, 80% of the extracted lenses had sodium concentrations in this intermediate range. The data indicate that the osmotic stresses induced in the lenses of diabetic and non-diabetic patients are different. The diabetic lenses were also distributed in the middle range when nuclear colour was graded on a scale from I to V, while normal lenses were again normally distributed at either end of the scale.