Spontaneous Spiking Activities of Porcine Pars intermedia Cells: Effects of Thyrotropin-Releasing Hormone

Abstract

The electrophysiological properties of porcine melanotrophs in primary culture were studied with patch clamp techniques. In the cell-attached (c/a) configuration, extracellular spikes were recorded but in only 47 of the 259 cells examined; active cells were more frequently found in short-term cultures (48 h, 22 out of 71). In the whole-cell (WCR) configuration, the mean resting potential was -46.6 mV and the mean input resistance 2.41 G omega. In the current clamp mode, of the 99 cells recorded, 39.4% displayed spontaneous spiking activities, 14.1% spiked when a depolarizing current was applied, and 46.5% were silent. Two patterns of spontaneous activity were recorded. The first consisted of rapid membrane depolarizations (mean duration: 16.0 ms) which reached a mean value of +8.9 mV. These action potentials occurred either at random intervals or at a mean frequency of 1.19 Hz. Tetrodotoxin (TTX), a sodium channel blocker, completely abolished these action potentials. The second pattern consisted of regular bursts (mean duration: 1.69 s and mean frequency: 0.15 Hz) of spikes with smaller amplitudes (culminating at -9.5 mV) and greater durations (79.7 ms). This pattern could be recorded in the presence of TTX in the external medium. In the c/a configuration, thyrotropin-releasing hormone (TRH) and (3Me-His2)-TRH enhanced spike frequency, whereas histidyl-proline-diketopiperazine, a metabolite of TRH, had no effect. In WCR, out of the 27 cells tested, TRH (5.10(-9)-5.10(-8) M) induced firing in 4 quiescent cells and increased the frequency of action potentials in 4 spontaneously active cells. This was usually, but not necessarily, preceded by a hyperpolarization (n = 5). TRH (5.10(-8) M) enhanced the secretion of melanocyte-stimulating hormone (alpha MSH) from perfused isolated melanotrophs by 62.8 +/- 9.8% (n = 9) over basal levels. (3Me-His2)-TRH and (Pro3)-TRH mimicked the TRH response, whereas histidyl-proline-diketopiperazine was without effect.