Effects of lutropin (LH) receptor internalization and recycling on the apparent numbers of LH receptors in rat testis Leydig cells at different temperatures

Abstract The effect of different temperatures on the Scatchard analysis of binding studies with 125 I-hCG and intact rat testis Leydig cells has been investigated. The results show that at temperatures greater than 4°C an overestimation of the number of receptors/cell is likely to occur. A one-site analysis of the data gives values of 13573, 61924 and 3802 LH receptors/cell at 34°C., 21°C and 4°C respectively. At 34°C and 21°C (but not at 4°C) a two-site analysis of the data is possible, giving similar high affinity binding components at both temperatures ( K D ∗ 1 × 10 −10 M ) but with dissimilar low affinity binding components (34°C, 2.17 ×X 10~9 M and 21°C., 6.3 X 10~8 M). The calculated total number of LH receptors using a two-site analysis at 21 °C is equal to 114766 receptors/cell, and at 34 °C it is 37987 receptors/cell. It is proposed that the differences in the level of binding at different temperatures and the associated changes in the value of K D , reflect the temperature sensitivity of the endocytic pathway of th 123 e 125 I-hCG/LH receptor complex within the rat testis Leydig cell. From previous studies it is known that at 34 °C and 21°C., a large part of the cell-associated hormone is not receptor bound but rather it represents ‘processed’ hormone before it is released from the cell. At 4°C the cell-associated 125 I-hCG remains bound only to cell surface LH receptors and thus gives a more accurate measure of total receptor numbers/cell.