Brain Research, 582 (1992) 357-358 (~) 1992 Elsevier Science Publishers B.V. All rights reserved. 0006-8993/92/$05.00

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BRES 25215

Effects of advancing age on cerebrospinal fluid concentrations of prolactin in the female rat James W. Simpkins Deparrnent of Pharmacodynamws and The Center for the Neurobiology of Agmg, Umverstty of Flortda, Gameswlle, FL 32610 (USA) Key words. Cerebrospinal fluid, Prolactm, Aging; Blood-brain barrier Studies were conducted to determine the effects of advancing age on serum and cerebrospinal fluid (CSF) prolactin (PRL) concentrations in the female rat. In young rats with basal serum PRL levels, CSF PRL was maintained at 1.1-2.1% of serum PRL levels In middle-aged rats, CSF PRL levels and the raUo of CSF to serum PRL was low in rats which maintained estrous cycles, but was increased 4- to 10-fold in rats which were in constant estrus. In aged constant estrous rats, CSF PRL concentrations were increased markedly and the CSF to serum PRL ratio increased to 31% Collectively, these data indicate that (i) in young female rats, only a fraction of serum PRL reaches the CSF; (n) that CSF PRL concentrations are low in middle-aged rats which exhibit estrous cycles and (iii) that aging and the constant estrous state are associated with elevated CSF PRL concentrations. As such, the age-related elevation in serum PRL levels and the fraction of PRL which accumulates m the CSF may contribute to the age-related dysfunction in brain PRL-responsive neurons. Prolactin (PRL) gains access to the brain and has b e e n quantitated in cerebrospinal fluid (CSF) 2'4. P R L activates t u b e r o i n f u n d i b u l a r d o p a m i n e ( T I D A ) n e u r o n s (see ref. 1 for references) and chronic hyperprolactinemia causes a disruption in T I D A function6'8'9 in a m a n n e r which simulates the age-related decline in T I D A function (see ref. 10 for references). As rats show an agerelated hyperprolactinemia (see ref. 10 for references), this endocrine condition may contribute to aging of T I D A neurons. The age-related damage to T I D A n e u r o n s and perhaps other brain D A n e u r o n a l systems could represent the cumulative effect of life-long exposure to normal P R L levels. Alternatively, if with age, P R L more readily penetrates the blood-brain-barrier (BBB), the aged rat may experience an exacerbated effect of P R L on D A n e u r o n a l function. The present study assessed the age-related changes in serum and CSF P R L concentrations in an effort to resolve this issue. L o n g - E v a n s female rats were purchased from Charles Rivers Labs (Wilmington, M A ) at 2 - 3 or 8-10 months of age. Animals were used in experiments when they were 3-4, 10-12 or 22 months of age. Estrous cycles were monitored daily for at least 10 days per m o n t h to establish the reproductive status of the animals. The young rats used exhibited 4- to 5-day estrous cycles (normally cycling, NC). The middle-aged rats exhibited 6- to 8-day estrous cycles (extended estrous; E E ) or constant estrous (CE) and the aged animals were all in CE. Animals from each age group were anesthetized with dimethyl ether and a single 1 ml blood sample was obtained by cardiac puncture for later assay of serum P R L

concentrations. Animals were maintained u n d e r ether anesthesia and placed in the ear bars of a David Kopf (Model 900) Steriotaxic instrument to secure the head in a position to expose the cisterna magna. CSF was collected from a single sample by exposure of the cisterna magna through an incision made at the base of the dorsal aspects of the skull. A 50-100/~1 sample of CSF was slowly withdrawn. A n y samples with evidence of blood in the CSF sample were discarded. All samples were obtained between 09.00 and 12.00 h. Serum P R L concentrations were determined in quadruplicate (replicates at 2 serum volumes) and CSF P R L TABLE I

Serum and CSF concentrattons of prolactm tn young (3-4 months), middle-aged (10-12 months) and old (22 months) LongEvans female rats PRL levels in ng/ml, n = 6-13 rats for Study 1 and 4-8 rats for Study 2. 1 All samples were below the limits of detectabdlty of the assay (0.3 ng/ml)

Study 1 Serum CSF Study 2 Serum CSF

Young

Mtddle-aged

Old

(NC)

(EE)

18 6+8.7 0.4+0.1

47+15 0.68+0.19

283+50c 67___32 2.2+0.8a'b 87_+2U

26.4+9 4 0 31

74+19 0.32+0.02

145+33a'b 4 9+2.4 a'b

(CE)

ap < 0.05 vs young NC. bp < 0.05 vs middle-aged EE cp < 0 05 vs all other groups.

Correspondence. J.W. Slmpkms, Department of Pharmacodynamlcs, University of Florida, Gainesville, FL 32610, USA.

(CE)

358 concentrations were d e t e r m i n e d from a single sample. CSF volumes used ranged from 50 to 100/~1. The R I A used reagent kindly provided by A . F. Parlow and the National Pituitary H o r m o n e Distribution Program of N I D D K . The reference p r e p a r a t i o n , rat P R L RP-1 was used to quantitate h o r m o n e concentrations. The intraassay coefficient of variation ranged from 4.3 to 5.7% for the assays used in these studies. One-way analysis of variance and S t u d e n t - N e w m a n Keul's tests were e m p l o y e d for all comparisons and a probability level of P < 0.05 was considered significant. Serum P R L concentrations were highly variable m young and middle-aged rats, a likely result of bleeding under ether anesthesia (Table I). A l t h o u g h m e a n P R L concentrations increased in middle-aged versus young rats, these differences were not significant (Table I). In aged CE rats serum P R L concentrations were elevated and were significantly higher than those observed in young and middle-aged animals. CSF P R L concentrations did not increase through middle-age if animals continued to exhibit estrous cycles (Table I). H o w e v e r , in middle-aged CE rats, CSF P R L levels were increased approximately 4-fold and in aged C E rats C S F P R L concentrations increased to 87 + 21 ng/ml, a value which was 31% of serum P R L concentrations. Similar data were obtained in a second study in which a significant increase in C S F P R L concentrations was observed, in association with a significant increase in serum P R L in middle-aged C E rats, but not in age-matched E E animals. In rats which maintained relatively normal estrous cycles, increasing age from 3 - 4 to 10-12 months had little effect on serum or CSF P R L concentrations. Thus, through middle-age in the rat, the maintenance of estrous cycles is associated with the maintenance of normal CSF P R L concentrations. H o w e v e r , in middle-aged rats which had been in the CE state for about 2 months, serum and CSF P R L concentrations were increased and the C S F to serum ratio of P R L was elevated. These data indicate that during middle-age, the brain ~s exposed to

4- to 10-fold m o r e P R L than the brains of ammals which maintain relatively normal estrous cycles. With increasing age and increasing length of time in the reproductive senescent C E state, the levels of serum P R L increase and the fraction of serum P R L in the CSF increases to as much as 31%. Thus, it appears that reproductive senescence in the rat places an even increased P R L load on the brain. This may contribute to neurodegenerative events, particularly in the T I D A systems, which show D A depletion during chronic hyperprolactinemia 6's'9. The mechanism of the enhanced fractional delivery of P R L to the CSF in aging CE rats is unknown, but two factors are apparent. First, female rats exhibit pituitary h y p e r t r o p h y with age 3 and many aged female rats exhibit anterior pituitary a d e n o m a s a'l°. In our study, anterior pituitary weights in young, middle-aged and aged rats were 12.3 + 0.8, 13.1 ___0.9 and 16.2 + 1.3, respectively. Thus, an age-related increase in the size of the pituitary could have contributed to the observed elevation in C S F P R L . Pituitary a d e n o m a s were not, however, a contributing factor since ammals showing evidence of unilateral pituitary h e m o r r h a g e were r e m o v e d from the study and their samples were discarded. Additionally, age-related changes in the BBB and/or the blood-cerebrospinal fluid barrier could contribute to the observed increased permeability to PRL. A n age-related d e c r e m e n t in the integrity of the BBB T M and of the b l o o d - c e r e b r o s p i n a l fluid barrier 7 has been described, with capillary endothelial cells exhibiting a b n o r m a l intercellular junctions in the BBB and the separation of normally closely juxtaposed tanycytes in the blood-cerebrospinal fluid barrier. Taken together, the elevation in serum P R L and the decline in integrity of the BBB and the blood-cerebrospinal fluid barrier could contribute to the age-related increase in CSF P R L observed in the present study. This set of circumstances could accelerate the rate of aging of PRL-responsive neurons in the brain.

1 Annunzlato, L , Regulation of the tuberomfundlbular and mgrostriatal systems, Neuroendocrmology, 29 (1979) 66-76 2 Clemens, J A. and Sawyer, B.D , Identification of prolactm m CSF, Bram Res., 21 (1974) 399-402. 3 Coleman, G.L., Barthold, S W., Osbaldiston, G , Foster, S J and Jonas, A M., Pathological changes during aging m barrier reared Fischer 344 male rats, J Gerontol, 32 (1977) 258-278 4 Logm, I S. and MacLeod, R.M, Prolactm in human and rat serum and cerebrosplnal fired, Brain Res., 132 (1977) 477-483. 5 Oztas, B , Kaya, M and Camurcu, S , Age related changes in the effects of electroconvulsive shock on the blood-brain-barrier permeability in rats, Mech Aging Dev . 51 (1990) 149-155 6 Sarkar, D K., Gottschall, P E and Meltes, J , Damage to hypothalamlc dopammerglc neurons Is associated with development of prolactln-secretmg pituitary tumors, Science, 218 (1982) 648-686

7 Scott. D E and Sladek, J R , Jr, Age-related changes m the endocnne hypothalamus I Tanocytes and the blood-brain-cerebrosplnal flmd barner, Neuroblol. Agmg, 2 (1981) 89-94 8 Stmpklns, J.W. and Gabnel, S.M., Chromc hyperprolactinemla causes progressive changes m hypothalamlc dopaminerglc and noradrenerglc neurons, Brain Res , 309 (1984) 277-282 9 Slmpklns, J W, Hodson, C A , Kalra, P.S. and Kalra, S.P., Chronic hyperprolactmemla depletes hypothalamlc dopamlne concentrations m male rats, Life Set , 30 (1982) 1349-1353. 10 Simpkins, J.W and Millard, W.J, Influence of age on neurotransmitter function In B. Sacktor (Ed.), Endocrmology and

The author wishes to thank B.J. Gores for her expert secretarial assistance and e&tonal review of the manuscript Supported by NIH Grant AG02021

Agmg, Endocrine and Metabohe Clinics of North Amerwa, Vol 16, Philadelphia, PA, 1987, pp. 893-917

11 Topple, A., Flfkova, E and Cullen-Dockstader, K , Effects of age on blood vessels and neurovascular apposmons in the rat dentate fascia, Neurobtol Aging, 11 (1990) 371-380.

Effects of advancing age on cerebrospinal fluid concentrations of prolactin in the female rat.

Studies were conducted to determine the effects of advancing age on serum and cerebrospinal fluid (CSF) prolactin (PRL) concentrations in the female r...
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