**7. Leptin regulation of somatotropes**

Somatotropes are vital metabolic sensors because they directly regulate stores of fat as they build muscle, bone, and regulate optimal body composition [63]. Most somatotropes bear leptin receptors [64, 65] and leptin deficiency results in reduced somatotrope functions [4, 20, 66–68]. As stated in the introduction, leptin treatment of leptin deficient *ob/ob* mice restores pituitary GH secretion and *Ghrhr* mRNA levels, but not hypothalamic production or secretion of GHRH [24].

Our studies of leptin's regulation of somatotropes began with the ablation of LEPR exon 17 or exon 1 with Cre-recombinase driven by the rat GH promoter [22, 69]. Both models showed GH deficiency, adult-onset obesity and metabolic dysfunction. At the level of the pituitary, this deficiency was seen as a reduction in GH and GHRHR.

We also reported sex-specific deficiencies during postnatal development with the discovery that leptin may target two transcription factors important in the production of GH, GHRHR, PRL, and TSH. These included Prophet of Pit1 (Prop1) and Pou1f1 [70]. Ablation of LEPR exon 1 in somatotropes reduced Pou1f1 in neonatal females along with serum prolactin. GH stores detected by immunolabeling were also reduced in both neonatal males and females. Interestingly, the lack of LEPR promoted an increase in Prop1 in neonatal males.

The studies of the impact of loss of LEPR were continued on FACS purified somatotropes [60]. Purified somatotropes showed reductions in GH, as expected, however they also contained a subset of multihormonal cells storing TSH and/or prolactin. In somatotrope LEPR-null females, TSH and prolactin stores were reduced in the pure somatotrope fraction [60]. Taken together, our analysis of somatotropes that lack LEPR shows that this multihormonal subset is significantly reduced, suggesting once more that leptin may play a role in maintaining multihormonal expression and promoting pituitary cell plasticity. Finally, these studies also demonstrated that Pou1f1 was reduced in pure somatotropes, which may explain the reduction in any or all hormones dependent on this transcription factor (GH, GHRHR, TSH and prolactin) [60].

We continued the investigation of leptin signaling pathways in somatotropes and reported that they included both transcriptional and posttranscriptional regulators [3]. Our tests of pathway inhibitors showed that full GH expression may be maintained by leptin through the JAK/STAT3 pathway but not nitric oxide. This contrasts with leptin pathways that regulate gonadotropins, which include NOS. Leptin regulation is likely to be transcriptional as loss of LEPR in somatotropes reduced *Gh* and *Ghrhr* mRNA and proteins [3]. In addition, leptin regulation of the *Pou1f1* transcription factor may also serve as a pathway for the transcriptional regulation of *Gh* and *Ghrhr* [2, 3, 60].

However, regulation of POU1F1 by leptin appears to be via post-transcriptional mechanisms as loss of LEPR in somatotropes causes reduction in mRNA levels of

the Pou1f1 protein, but not the *Pou1f1* mRNA [2, 60]. Conversely, leptin stimulation results in increased expression of Pou1f1 proteins, but not mRNA [60].

An *in silico* analysis detected eight Musashi binding elements in the 3'UTR of the *Pou1f1* mRNA and tests of Musashi binding showed direct interaction of Musashi with this region and repression of translation, which was reversed by leptin [2]. Furthermore, Musashi immunoprecipitation of whole pituitary extract showed coassociation of Musashi and the endogenous *Pou1f1* mRNA. Our analyses of transcripts by scRNA-sequencing studies of normal pituitary cells showed that *Msi1* mRNA was expressed in somatotropes. This was confirmed in pure somatotrope populations [2].
