Trace Nutrients Research22:101-106(2005)

Trace Nutrients Research 22 : 101 − 106（2005）

Excessive accumulation of calcium and movement of parathyroid hormone (PTH) inthe olfactory system by zinc deficiency
Takao KURIYAMA1）, Kenichi TOMIYAMA1）, Moritoshi NIHIRA1）, Tomomi JYOHO1）, Takayuki TAKEUCHI2）,Yukihiro NAKANO2）, Harunobu NAKASHIMA3）, Yoshikazu MATSUDA4）and Yasuaki ARAKAWA1）
1）

Department of Hygiene & Preventive Medicine, Faculty of Health Sciences, The University of Shizuoka＊
Research Reactor Institute, Kyoto University＊＊2）

Osaka Prefectural Institute of Public Health＊＊＊3）

Central Research Institute, Japan Clinic Co., Ltd.＊＊＊＊

4）

Summary
Zinc deficiency induced an excessive accumulation of calcium in the olfactory system. This calcium localization
was parallel with the degree of symptom manifestation of zinc deficiency. Therefore, to investigate the mechanismon the calcium localization in the olfactory system, the movement of parathyroid hormone (PTH), cAMP and inositol
triphosphate (IP3) which play a part in olfactory signal transduction and Ca-homeostasis were analyzed in the olfactory epithelium, olfactory bulb and cerebrospinal fluid of zinc deficient rats.
Results were as follows; 1) Calcium showed a remarkable decrease in the cerebrospinal fluid and a remarkableincrease in the olfactory tissues. 2) Magnesium also showed a decrease in the cerebrospinal fluid and increases in
the olfactory tissues, although the movements were much lower than those of calcium. 3) PTH showed significant
increases in both the cerebrospinal fluid and the olfactory tissues at the progress period. However, significant
decreases in the olfactory tissues and a remarkable increase in the cerebrospinal fluid were seen at the perfection
period of zinc-deficiency. 4) cAMP revealed excessive increases in the olfactory epithelium and olfactory bulb at theprogress period of zinc deficiency. On the other hand, a little decreases in the olfactory epithelium and olfactory
bulb were seen at the perfection period of zinc-deficiency. 5) Regarding IP3, as well as cAMP, excessive increases in
the olfactory epithelium and olfactory bulb were seen at the progress period of zinc deficiency. At the perfection
period of zinc-deficiency, however, significant decreases were seen in the olfactory tissues.

Deficiency or excess of trace elements induce various kinds of brain lesion

1− 7）. In this study, zinc deficiency-induced olfactory lesions (anosmia) were kinetically analyzed from the viewpoints of the movement of trace elements, an inhibition of signal transduction and toxic cell death (necrosis or apoptosis).
In a series of this study, we found remarkable results that zinc deficiency induced an excessive accumulation of8−11）

calcium in the olfactory system such as olfactory epithelium and olfactory bulb

. This calcium localization wasparallel with the degree of symptom manifestation of zinc deficiency. Therefore, to investigate the mechanism on
the calcium localization in the olfactory system, the movement of parathyroid hormone (PTH), cAMP and inositol
triphosphate (IP3) which play a part in olfactory signal transduction and Ca-homeostasis were analyzed in the olfac＊＊＊＊Address：52−1, Yada, Suruga-ku, Shizuoka 422−8526, Japan＊＊＊＊Address：Kumotori-cho, Sennan-gun, Osaka 590−0494, Japan＊＊＊＊Address：1−3−69, Nakamichi, Higashinari-ku, Osaka 537−0025, Japan＊＊＊＊Address：10−1, Kainichi-cho, Uzumasa, Ukyo-ku, Kyoto 616−8555, Japan― 101 ―

Trace Nutrients Research22:101-106(2005)

tory epithelium, olfactory bulb and cerebrospinal fluid of zinc deficient rats.

Materials and Methods
The animals used in this study were 3 weeks old, SPF Wistar-derived male rats purchased from Japan SLC Inc.
(Hamamatsu, Japan). Rats fed zinc-deficient diet (Zn content: 0.05 mg%, Oriental Yeast Co. Ltd) or zinc-supplement
diet (control diet, ZnCO3 added, Zn content: 5.8 mg%, Oriental Yeast Co. Ltd) for 1 week or 4 weeks.
Olfactory epithelium, olfactory bulb and cerebrospinal fluid were collected from rats fed zinc-deficient diet or zincsupplement diet for 1 week (zinc deficient progress period) or 4 weeks (zinc deficient perfection period) after thestarting of feeding.
Concentration of Ca and Mg in the olfactory epithelium, olfactory bulb and cerebrospinal fluid were analyzed by12）
using a thermal neutron activation analysis method .

PTH in the olfactory epithelium, olfactory bulb and cerebrospinal fluid were assayed with a PTH[125I] radio13）
immuno assay .

cAMP in the olfactory epithelium and olfactory bulb were assayed with a cAMP[125I] radio immuno assay14）.

315）
IP3 in the olfactory epithelium and olfactory bulb were assayed with a IP3 [ H] radio receptor assay .

Results
Figure 1 shows the relationship of calcium, magnesium and PTH in the olfactory epithelium, olfactory bulb andthe cerebrospinal fluid of zinc-deficient rats. Concentraion of calcium showed a remarkable decrease in the cerebrospinal fluid and remarkable increases in the olfactory tissues. Regarding magnesium, a decrease in the cerebrospinal fluid and increases in the olfactory tissues were seen as well as calcium, although the movements weremuch lower than those of calcium. PTH levels showed significant increases in both the cerebrospinal fluid and theolfactory tissues at the progress period. However, significant decreases in the olfactory tissues and a remarkableincrease in the cerebrospinal fluid were seen at the completed stage of zinc-deficient manifestation.
Figure 2 shows the movement of cAMP and IP3 in the olfactory epithelium and the olfactory bulb of zinc-deficientrats. Concentraion of cAMP showed excessive increases in the olfactory epithelium and olfactory bulb at the progressive stage of zinc deficiency manifestation. On the other hand, a little decrease in the olfactory epithelium andolfactory bulb were seen at the completed stage of zinc-deficient manifestation. Regarding IP3, as well as cAMP,
excessive increases in the olfactory epithelium and olfactory bulb were seen at the early stage of zinc deficiency
manifestation. At the completed stage of zinc-deficient manifestation, however, significant decreases were seen inthe olfactory tissues.

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Trace Nutrients Research22:101-106(2005)

Fig. 1 The relationship of calcium, magnesium and parathyroid hormone(PTH) in the olfactory
epithelium, the olfactory bulb and the cerebrospinal fluid of zinc-deficient rats. SPF Wistarderived male rats (3 week ages) fed with zinc-deficient diet or zinc-supplement diet (controldiet) for 1 (progress period) or 4 weeks (perfection period).
Vertical bars in the mean of SD; those marked with asterisks differ significantly from the corresponding control value. ＊p<0.05 ＊＊p<0.01 ＊＊＊p<0.001― 103 ―

Trace Nutrients Research22:101-106(2005)

Fig. 2 Movement of cAMP and inositol-1,4,5-triphosphate (IP3) in the olfactory epithelium and theolfactory bulb of zinc-deficient rats. SPF Wistar-derived male rats (3 week ages) fed withzinc-deficient diet or zinc-supplement diet (control diet) for 1 (progress period) or 4 weeks(perfection period).

Discussion
Concentration of calcium showed a remarkable decrease in the cerebrospinal fluid and a remarkable increase in2+

the olfactory tissues. This means an excessive influx of extracellular Cafrom the cerebrospinal fluid to the olfacto-ry tissues.
Content of magnesium showed a decrease in the cerebrospinal fluid, but a little increase in the olfactory tissues.
This result revealed that influx of extracellular Mg

2+from the cerebrospinal fluid to the olfactory tissues wasextremely small.
An increase and a decrease of calcium and magnesium are known as hypercalcemia and hypermagnesemia, whichwere regulated by PTH. Hypercalcemia and hypermagnesemia were recognized in the organs other than the brain.
No reports showed PTH production in the brain except for a report referring to PTH production in the brain of16）
sheep . We speculated that the excessive accumulation of calcium in the olfactory tissues may be related to PTH in

the brain. Our experimental results showed that a significant increases of PTH were seen in both the cerebrospinal
fluid and the olfactory tissues, followed by the activation of adenylcyclase by the excessive increases of PTH in thespecial regions of the brain, excessive increases in the formation of cAMP by the activation of adenylcyclase in the― 104 ―Trace Nutrients Research22:101-106(2005)

early stage (progressive stage) of symptom manifestation of zinc deficiency. These results mean an excessive accel2+
eration of extracellular Ca -influx to the tissues. At the completed stage of zinc-deficient manifestation, however,significant decreases of PTH in the olfactory tissues and a remarkable increase of PTH in the cerebrospinal fluid
were seen. These significant decreases of PTH in the olfactory tissues appear to be due to the decreases in the affinity and bonding amount of PTH to the receptor which are caused by the break-down of metabolic systems and cell
death under the long-term abnormal condition. Conversely, a remarkable increase of PTH in the cerebrospinal fluid
appear to be due to an excessive acceleration of PTH generation to recovery the low levels of calcium and magnesium in the cerebrospinal fluid and significant decreases of PTH in the olfactory tissues.
cAMP increased in the olfactory epithelium and olfactory bulb at the progressive stage of zinc deficiency manifestation. This means an excessive acceleration of cAMP driving processes such as the activation of adenylcyclase by
the excessive increases of PTH at Ca channel. On the other hand, a little decreases of cAMP in the olfactory epithelium and olfactory bulb were seen at the completed stage of zinc-deficient manifestation. These decreases in the tissues appear to be due to the working of negative feed-back, the break-down of metabolic systems and cell deathunder the long-term abnormal condition. We observed extensive destruction of the cortical neurons at the completed stage of zinc-deficient manifestation (data are not shown).
Regarding IP3, as well as cAMP, excessive increases of IP3 in the olfactory epithelium and olfactory bulb wereseen at the early stage of zinc deficiency manifestation. This means an excessive acceleration of IP3 working
2+processes such as IP3 -dependent Ca

channel and intracellular Ca2+-mobilization. At the completed stage of zinc-

deficient manifestation, however, remarkable decreases of IP3 were seen in the olfactory tissues. These remarkabledecreases in the tissues appear to be due to the working of negative feed-back, the break-down of metabolic systems and cell death under the long-term abnormal condition, as described above.
Putting all results together, the calcium localization in the olfactory system seems to be due to the processes suchas the activation of adenylcyclase by an excessive increase of PTH in the special regions of the brain, excessive
increase in the formation of cAMP by the activation of adenylcyclase, disorder of cAMP-activated channels by the2+excessive increase of cAMP, and an excessive influx of extracellular Ca

by the disorder of the channels. Moreover,this rapid and excessive increase in the concentration of intracellular Ca2+

seems to inhibit CaM-kinase II functions

and to activate endonuclease, followed by leading to toxic cell death (necrosis or apoptosis).

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