Allopregnanolone modulates striatal dopamingergic activity of rats under different gonadal hormones conditions.

OBJECTIVES: Progesterone modulates dopamine (DA) release in corpus striatum. Our objective was to evaluate the effect of the i.c.v injection of the neurosteroid allopregnanolone (ALL), a progesterone metabolite on dopaminergic activity in the corpus striatum of rats under different gonadal hormonal conditions. METHODS: We have measured the concentrations of DOPA, DA and DOPAC (main metabolite of DA) in the corpus striatum in estrus and diestrus rats and in ovariectomized rats without hormonal replacement (OVX group) and primed with estrogen and progesterone (OVX(i) group). Additionally, we have used the aromatic acid decarboxylase inhibitor NSD in order to evaluate the function of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis. RESULTS: ALL significantly decreased the striatal concentrations of both DA and DOPAC in the estrus. On the other hand, ALL increased significantly the levels of DA in the OVX(i) group. The DOPA accumulation in OVX(i) after NSD treatment in the ALL-treated groups was greater than in the vehicle group. However, the estrus group did not modify the DOPA accumulation after NSD injection. DISCUSSION: Our results suggest that ALL could modulate the dopaminergic transmission in the corpus striatum by causing changes in the activity of TH and/or in the pre- and post-synaptic dopaminergic terminals in the corpus striatum. This neurosteroidal mechanism could be a new kind of neurotransmitter systems modulation accomplished on TH activity itself and/or on the second messengers not related to ionic channels. Additionally, our results reinforce the idea of a close relationship between the fast non-genomic mechanism of ALL and the genomic actions of estrogen and progesterone.

Gene therapy in the neuroendocrine system.

The implementation of experimental gene therapy in animal models of neuroendocrine diseases is an area of growing interest. In the hypothalamus, restorative gene therapy has been successfully implemented in Brattleboro rats, an arginine vasopressin (AVP) mutant which suffers from diabetes insipidus, and in Koletsky (fa(k)/fa(k)) and in Zucker (fa/fa) rats which have leptin receptor mutations that render them obese, hyperphagic and hyperinsulinemic. In the above models, viral vectors expressing AVP, leptin receptor b and proopiomelanocortin, respectively, were stereotaxically injected in the relevant hypothalamic regions. In rats, aging brings about a progressive degeneration and loss of hypothalamic tuberoinfundibular dopaminergic (TIDA) neurons, which are involved in the tonic inhibitory control of prolactin secretion and lactotropic cell proliferation. Stereotaxic injection of an adenoviral vector expressing insulin-like growth factor I corrected their chronic hyperprolactinemia and restored TIDA neuron numbers. Spontaneous intermediate lobe pituitary tumors in a retinoblastoma (Rb) gene mutant mouse were corrected by injection of an adenoviral vector expressing the human Rb cDNA and experimental prolactinomas in rats were partially reduced by intrapituitary injection of an adenoviral vector expressing the HSV1-thymidine kinase suicide gene. These results suggest that further implementation of gene therapy strategies in neuroendocrine models may be highly rewarding.

Gene therapy for long-term restoration of circulating thymulin in thymectomized mice and rats.

Thymulin is a thymic peptide possessing hypophysiotropic activity and antiinflammatory effects in the brain. We constructed a synthetic DNA sequence encoding met-FTS, a biologically active analog of thymulin, and subsequently cloned it into different expression vectors. A sequence optimized for expression of met-FTS in rodents, 5'-ATGCAGGCCAAGTCGCAGGGGGGGTCGAACTAGTAG-3', was cloned in the mammalian expression vectors pCDNA3.1(+) and phMGFP (which expresses the Monster Green Fluorescent Protein), thus obtaining pcDNA3.1-metFTS and p-metFTS-hMGFP, which express met-FTS and the fluorescent fusion protein metFTS-hMGFP, respectively. The synthetic sequence was also used to construct the adenoviral vector RAd-metFTS, which expresses met-FTS. Transfection of HEK293 and BHK cells with pcDNA3.1-metFTS (experimental groups) or pcDNA3.1 (control), led to high levels of thymulin bioactivity (>600 versus <0.1 pg/ml in experimental and control supernatants, respectively). Transfection of HEK293 and BHK cells with pmetFTS-hMGFP revealed a cytoplasmic and nuclear distribution of the fluorescent fusion protein. A single intramuscular (i.m.) injection (10(7) plaque forming units (PFU)/mouse or 10(8) PFU/rat) of RAd-metFTS in thymectomized animals (nondetectable serum thymulin) restored serum thymulin levels for at least 110 and 130 days post-injection in mice and rats, respectively. We conclude that RAd-metFTS constitutes a suitable biotechnological tool for the implementation of thymulin gene therapy in animal models of chronic brain inflammation.