Tilapia prolactin cells are thermosensitive osmoreceptors

Abstract

Prolactin (PRL) cells within the rostral pars distalis (RPD) of euryhaline and eurythermal Mozambique tilapia, Oreochromis mossambicus, rapidly respond to a hyposmotic stimulus by releasing two distinct PRL isoforms, PRL₁₈₈ and PRL₁₇₇. Here, we describe how environmentally relevant temperature changes affected mRNA levels of prl₁₈₈ and prl₁₇₇ and the release of immunoreactive prolactins from RPDs and dispersed PRL cells. When applied under isosmotic conditions (330 mosmol/kgH₂O), a 6°C rise in temperature stimulated the release of PRL₁₈₈ and PRL₁₇₇ from both RPDs and dispersed PRL cells under perifusion. When exposed to this same change in temperature, ∼50% of dispersed PRL cells gradually increased in volume by ∼8%, a response partially inhibited by the water channel blocker, mercuric chloride. Following their response to increased temperature, PRL cells remained responsive to a hyposmotic stimulus (280 mosmol/kgH₂O). The mRNA expression of transient potential vanilloid 4, a Ca²⁺-channel involved in hyposmotically induced PRL release, was elevated in response to a rise in temperature in dispersed PRL cells and RPDs at 6 and 24 h, respectively; prl₁₈₈ and prl₁₇₇ mRNAs were unaffected. Our findings indicate that thermosensitive PRL release is mediated, at least partially, through a cell-volume-dependent pathway similar to how osmoreceptive PRL release is achieved.

Keywords: fish; osmoreceptor; prolactin; thermoreceptor; thermosensitivity.

Graphical abstract

Figure 1.

Diagram summarizing the experimental setup used for determining changes in PRL cell volume in responses to a rise in temperature. PRL, prolactin.

Figure 2.

Effects of temperature on the release of PRL₁₈₈ (A) and PRL₁₇₇ (B) from RPDs at 1, 6, and 24 h and on the release of PRL₁₈₈ (C) and PRL₁₇₇ (D) from PRL cells at 1 and 6 h. A and B: data are expressed in ng/mL/g body wt ± SE (n = 7 or 8). Effects of incubation time and temperature were analyzed by two-way ANOVA (***P < 0.001). Incubation temperature effects were followed up by protected Fisher’s LSD test. Symbols not sharing the same letter are significantly different across temperatures at P < 0.05. C and D: data are expressed in µg/10⁵ cells ± SE (n = 7 or 8). Effects of incubation time and temperature were analyzed by Student’s t test at each time point (*P < 0.05). LSD, least-significant different; PRL, prolactin; RPDs, rostral pars distalis.

Figure 3.

Effects of temperature increase (from 26°C to 32°C) and reduction in osmolality (from 330 to 280 mosmol/kgH₂O) on the release of PRL₁₈₈ (A) and PRL₁₇₇ (B) from dispersed PRL cells within 105 min. Vertical lines indicate a change in temperature or osmotic conditions. Symbols represent mean percent change PRL release ± SE (n = 5 or 6). Treatment and time effects were analyzed by two-way ANOVA (***P < 0.001; **P < 0.01) followed by protected Fisher’s LSD test. Means with daggers (†) indicate significant differences from parallel controls at each time point (†P < 0.05, ††P <0.01). Control means with double daggers (‡) indicate differences from control time 0 (Fisher’s protected LSD; ‡P < 0.05, ‡‡P < 0.01, ‡‡‡P < 0.001). Temperature means with silcrows (§) indicate differences from temperature time 0 (Fisher’s protected LSD; §P < 0.05, §§P < 0.01, §§§P < 0.001). LSD, least-significant different; PRL, prolactin.

Figure 4.

Effects of temperature and osmolality on the change in volume (A) and NCCV (B) of dispersed PRL cells within 175 min. A: symbols represent means expressed as percent change from an average cell volume of the first 6 time points ± SE (n = 10–12). Vertical lines indicate a change in treatment. B: the bars represent NCCV during control (26°C, 330 mosmol/kgH₂O), temperature (32°C, 330 mosmol/kgH₂O), recovery (26°C, 330 mosmol/kgH₂O), and hyposmotic treatments (26°C, 280 mosmol/kgH₂O) as a % change from the baseline. White, black, and gray bars represent control, temperature, and temperature + HgCl₂ experimental groups, respectively. Effects of treatment and experimental group were analyzed by two-way ANOVA (***P < 0.001), followed by protected Fisher’s LSD test. Means not sharing the same letter (lowercase, between treatments; uppercase, between experimental groups) are significantly different at P < 0.05. LSD, least-significant different; NCCV, net change in cell volume; PRL, prolactin.

Figure 5.

Effects of temperature on the mRNA expression of prl₁₈₈ (A), prl₁₇₇ (B), and trpv4 (C) in RPDs at 24 h and prl₁₈₈ (D), prl₁₇₇ (E), and trpv4 (F) in PRL cells at 6 h. RPDs and PRL cells were incubated in 330 mosmol/kgH₂O media in temperatures ranging from 20°C to 32°C. A–C: data are expressed as fold-change from 20°C ± SE (n = 6–8) and analyzed by one-way ANOVA followed by protected Fisher’s LSD test. Bars not sharing the same letters are significantly different at P < 0.05. D–F: data are expressed as fold-change from the 26°C group ± SE (n = 7 or 8) and analyzed by Student’s t test (*P < 0.05). LSD, least-significant different; PRL, prolactin; RPDs, rostral pars distalis.