Lead (Pb2+) and cadmium (Cd2+) inhibit the dipsogenic action of central beta-adrenergic stimulation by isoproterenol

Braz J Med Biol Res, March 1997, Volume 30(3) 419-423 (Short Communication)

Lead (Pb ²⁺ ) and cadmium (Cd ²⁺ ) inhibit the dipsogenic action of central beta-adrenergic stimulation by isoproterenol

J.B. Fregoneze¹, C.A. Marinho², T. Soares¹, L. Castro¹, C. Sarmento², M. Cunha², V. Gonzalez², P. Oliveira¹, T. Nascimento², C.P. Luz¹, P. Santana Jr.², I.R. De-Oliveira² and E. De-Castro-e-Silva²

¹Departamento de Zoologia, Instituto de Biologia, Universidade Federal da Bahia, 40170-110 Salvador, BA, Brasil

²Departamento de Fisiologia, Instituto de Ciências da Saúde, Universidade Federal da Bahia, 40110-100 Salvador, BA, Brasil

^Text

^{Correspondence and Footnotes}

Abstract

We have previously demonstrated that acute third ventricle injections of both Pb²⁺ and Cd²⁺ impair the dipsogenic response elicited by three different situations: dehydration and central cholinergic or angiotensinergic stimulation. ß-Adrenergic activation is part of the multifactorial integrated systems operating in drinking behavior control in the central nervous system. In the present study acute third ventricle injections of Pb²⁺ (3, 30 and 300 pmol/rat) or Cd²⁺ (0.3, 3 and 30 pmol/rat) blocked the dipsogenic response induced by third ventricle injections of isoproterenol (ISO; 160 nmol/rat) in a dose-dependent manner. Normohydrated animals receiving ISO + NaAc (sodium acetate) or saline (controls) displayed a high water intake after 120 min (ISO + saline = 5.78 ± 0.54 ml/100 g; ISO + NaAc = 6.00 ± 0.6 ml/100 g). After the same period, animals receiving ISO but pretreated with PbAc at the highest dose employed (300 pmol/rat) drank 0.78 ± 0.23 ml/100 g while those receiving ISO and pretreated with the highest dose of CdCl₂ (30 pmol/rat) presented a water intake of 0.7 ± 0.30 ml/100 g. Third ventricle injections of CdCl₂ (3 nmol/rat) or PbAc (3 nmol/rat) did not modify food intake in rats deprived of food for 24 h. Thus, general central nervous system depression explaining the antidipsogenic action of the metals can be safely excluded. It is concluded that both Pb²⁺ and Cd²⁺ inhibit water intake induced by central ß-adrenergic stimulation.

Key words: lead, cadmium, water intake, isoproterenol

Human exposure to heavy metals such as cadmium (Cd²⁺) and lead (Pb²⁺) may bring about several adverse reactions. Neurotoxicity induced by heavy metals is well-documented in humans and experimental animals (1). Both Cd²⁺ and Pb²⁺ reach the central nervous system either by crossing the blood-brain barrier or via retrograde axonal transport (2,3).

The presence of heavy metals in the central nervous system disrupts the functional integrity of several neurotransmitter pathways. Many independent reports confirm alterations in brain monoaminergic receptors and uptake after Pb²⁺ (4) and Cd²⁺ (5,6) intoxication.

We have recently used a new approach to study the acute actions of heavy metals on the central nervous system by analyzing the effects of third ventricle injections of very small amounts of Cd²⁺ or Pb²⁺ on water intake, a very easy and simple parameter to measure in rats. Using this methodology we demonstrated that acute intracerebroventricular (icv) injections of lead acetate (7) or cadmium chloride (8) block the dipsogenic response induced by three different situations, i.e., water deprivation and central angiotensinergic or cholinergic stimulation, in a dose-dependent manner. In the particular case of Cd²⁺, its antidipsogenic effect seems to be also due to a serotonergic activation of central 5-HT₂ receptors (8).

Brain control of thirst is achieved by a complex integrated network of neurotransmitters acting in concert. It is generally accepted that central catecholaminergic pathways exert a dual role on drinking behavior regulation, since a-adrenergic stimulation decreases (9) and ß-adrenergic (10,11) stimulation enhances water intake. However, more detailed studies have suggested that a-adrenergic stimulation may yield dipsogenic or antidipsogenic responses depending on the area studied and the doses employed.

The aim of the present study was to determine if the dipsogenic response elicited by central ß-adrenergic stimulation by isoproterenol (ISO) could be blocked by acute icv injections of cadmium chloride or lead acetate.

Adult male Wistar rats kept under controlled light and temperature conditions (lights on from 6:00 to 20:00 h, 26 ± 2^oC) were used in the experiments. The animals were implanted with a cannula into the third ventricle as described elsewhere (12) 5 days before the experimental sessions. The following substances were used: lead acetate (PbAc), cadmium chloride (CdCl₂ ) and isoproterenol (Sigma Chemical Co., St. Louis, MO). All substances were dissolved in saline solution. Microinjections were made using a 10-µl Hamilton syringe connected to a Mizzy-Slide-Pak needle (12 x 27 gauge) through a polyethylene extension (PE10). The volume injected was 2 µl during a period of 90 s. The experiments were always performed between 8:00 and 11:00 a.m. After sacrifice a third ventricle injection of a vital dye was used for the macroscopic localization of the cannula. Data from only those animals whose cannulae were correctly placed into the third ventricle were considered.

Normohydrated animals received third ventricle injections of ISO at a dose of 160 nmol/rat or saline. To study the effects of the heavy metals on isoproterenol-induced dipsogenic response, distinct groups of animals were pretreated with CdCl₂ (0.3, 3.0 and 30.0 pmol/rat) or PbAc (3, 30 and 300 pmol/rat) 45 min before ISO injections. Control animals compared to PbAc-treated groups received icv injections of NaAc, whereas control animals compared to CdCl₂-treated groups received icv injections of saline. We have previously demonstrated no significant alterations in water intake after icv injections of saline or NaAc.

To test if the antidipsogenic effect of Cd²⁺ and Pb²⁺ were not simply due to a general central nervous system depression, we briefly determined the effect of these metals on food intake. After 24 h of food deprivation, the animals received icv injections of CdCl₂ (control group receiving saline) or PbAc (control group receiving NaAc). Immediately after the injections, the animals were allowed to enter the food compartment within the metabolic cage. The food consumption of a powder chow was monitored during the same time intervals as used to record water intake. The amount of chow dispersed on the floor of the cage was taken into account both in control and experimental animals.

We used a computer software (GBSTAT, Dynamic Microsystems Inc., Silver Spring, MD) that performs the repeated measures analysis of variance (ANOVA) followed by the Scheffé test. Differences were considered to be significant when P<0.01. The cumulative water intake is reported as ml/100 g body weight (mean ± SEM).

As shown in Figure 1 (panel A), normohydrated animals receiving NaAc exhibited a very low water intake while those receiving NaAc 45 min prior to ISO (160 nmol/rat) exhibited a significant increase in drinking behavior. Pretreatment with PbAc (3, 30 and 300 pmol/rat) reduced the dipsogenic effect of ISO injections in a dose-dependent fashion.

Figure 1
- Effect of PbAc and CdCl₂ on water intake induced by third ventricle ISO injections. Data are reported as cumulative water intake (ml/100 g body weight). Panel A, Water intake in animals receiving NaAc (300 pmol/rat) + saline (open squares); NaAc (300 pmol/rat) + ISO (160 nmol/rat) (open circles); PbAc (3 pmol/rat) + ISO (160 nmol/rat) (filled triangles); PbAc (30 pmol/rat) + ISO (160 nmol/rat) (filled inverted triangles); PbAc (300 pmol/rat) + ISO (160 nmol/rat) (filled lozenges). Data are reported as mean ± SEM for 12 animals in each group. *P<0.01 compared to the respective NaAc + ISO controls (repeated measures ANOVA followed by the Scheffé test). Panel B, Water intake in animals receiving saline + saline (open squares); saline + ISO (160 nmol/rat) (open circles); CdCl₂ (0.3 pmol/rat) + ISO (160 nmol/rat) (filled circles); CdCl₂ (3.0 pmol/rat) + ISO (160 nmol/rat) (filled triangles); CdCl₂ (30.0 pmol/rat) + ISO (160 nmol/rat) (filled inverted triangles). Data are reported as mean ± SEM for 12 animals in each group. *P<0.01 compared to the respective saline + ISO controls (repeated measures ANOVA followed by the Scheffé test).

Figure 1 (panel B) shows the result of CdCl₂ injections on water intake induced by icv ISO injections. As in the previous experiment, normohydrated rats treated with saline drank very small amounts of water. ISO (160 nmol/rat) induced a powerful increase in drinking behavior. The central administration of CdCl₂ (0.3, 3.0 and 30 pmol/rat) reduced the dipsogenic response elicited by ISO injections in a dose-dependent manner.

Table 1 summarizes food intake data for fasted (24 h) animals treated with CdCl₂ (3 nmol/rat; controls receiving saline) or PbAc (3 nmol/rat; controls receiving NaAc). There were no significant statistical differences between groups treated with the metals and their respective controls.

The data presented here clearly demonstrate that both Cd²⁺ and Pb²⁺ acutely injected into the third ventricle of male rats impair the dipsogenic response elicited by the central administration of ISO. This response was not related to a general depressive status of the central nervous system after the injection of the metals, since food intake after fasting was normally maintained in animals receiving icv injections of Cd²⁺ or Pb²⁺. In addition, animals receiving Pb²⁺ or Cd²⁺ at the doses used here did not present motor disturbances that could explain the reduced water intake. Circling, hyperactivity, or sickness-like postures or behaviors were not observed.

It is well established that the presence of heavy metals in the central nervous system hampers the function of several neurotransmitter pathways. Recent reports have shown that Pb²⁺ selectively reduced muscarinic receptors and cholineacetyltransferase in some brain areas (13). Dopamine concentration and synthesis regulation seem to be altered by Pb²⁺ (14). Pb²⁺ at extremely low concentrations affects the glutamatergic system in the brain. Glutamate synthesis is significantly reduced and NMDA-evoked currents are inhibited by Pb²⁺ (15). After postnatal Pb²⁺ exposure noradrenaline levels are increased in many brain regions (16) while the noradrenergic turnover rate may be decreased (17).

Cd²⁺ is also able to alter the function of many brain neurotransmitters. Catecholaminergic and serotonergic transmission in the central nervous system may be affected by heavy metal exposure, with both stimulation and inhibition being reported by different researchers (1). Also, the level and distribution of several biogenic amines in the central nervous system are modified by Cd²⁺ (5).

We have recently reported that acute injections of PbAc (7) or CdCl₂ (8) into the third ventricle of rats block the dipsogenic response induced by three different situations, i.e., water deprivation and central cholinergic or angiotensinergic stimulation. The data now included in the present paper clearly show that both Cd²⁺ and Pb²⁺ are also capable of decreasing water intake following central ß-adrenergic stimulation.

Catecholaminergic control of drinking behavior is well documented. Indeed, periventricular noradrenergic systems are crucial for angiotensin-induced water intake (18) and selective destruction of discrete noradrenergic areas by the neurotoxic agent 6-hydroxydopamine reduces drinking (19). Central ß-adrenergic stimulation is an undisputable stimulus generating water intake (10,11).

The data presented here show that the previous third ventricle injection of minute amounts of Cd²⁺ or Pb²⁺ impairs the dipsogenic effect of central ß-adrenergic stimulation by ISO. So it is clear that the presence of these metals in the central nervous system disrupts the capacity of ISO to induce water intake. However, we cannot rule out the possibility that both Cd²⁺ and Pb²⁺ may inhibit a commom mechanism activated by angiotensin II, noradrenaline or acetylcholine.

The effects of the metals observed here are probably due to very fast biochemical effects in the central nervous system which may be related, at least in part, to derangement of calcium-mediated cellular processes. A challenge that lies ahead will be to exploit the relationship between calcium metabolism in the brain in response to acute heavy metal injections, and the effects on drinking behavior observed here. It is important to note that CdCl₂ was much more potent than PbAc in blocking the isoproterenol-induced dipsogenic response.

References

1. Sutoo D (1994). Disturbances in brain function by exogenous cadmium. In: Isaacson RL & Jensen KF (Editors), The Vulnerable Brain and Environmental Risks. Vol. 3. Toxins in Air and Water. Plenum Press, New York, 281-300.

2. Bradbury MWB & Deane R (1993). Permeability of the blood-brain barrier to lead. Neurotoxicology, 14: 131-136.

3. Arvidson B (1994). A review of axonal transport of metals. Toxicology, 88: 1-14.

4. Luthman J, Lidqvist E, Gerhardt GA, Olson L & Hoffer BH (1994). Alterations in central monoamine systems after postnatal lead acetate treatment in rats. Environmental Research, 65: 100-118.

5. Sutoo D, Akiyama K & Maeda I (1987). The effect of cadmium administration on the biogenic amine distributions and amounts in the mouse brain. Kitasato Archives of Experimental Medicine, 60: 157-166.

6. Rastogi RB, Merali Z & Singhal RI (1977). Cadmium alters behaviour and the biosynthetic capacity for catecholamines and serotonin in neonatal rat brain. Journal of Neurochemistry, 28: 789-794.

7. Fregoneze JB, Cunha M, Bulcão C, Ferreira H & De Castro e Silva E (1994). Acute effect of intracerebroventricular administration of lead on the drinking behavior of rats induced by dehydration or central cholinergic and angiotensinergic stimulation. Physiology and Behavior, 56: 129-133.

8. De Castro e Silva E, Ferreira H, Cunha M, Bulcão C, Sarmento C, De Oliveira IR & Fregoneze JB (1996). Effect of central acute administration of cadmium on drinking behavior. Pharmacology, Biochemistry and Behavior, 53: 687-693.

9. Callera JC, Saad WA, Camargo LAA, Renzi A, De Luca Jr LA & Menani JV (1994). Role of adrenergic pathways of the lateral hypothalamus on water intake and pressor response induced by the cholinergic activation of the medial septal area in rats. Neuroscience Letters, 167: 153-155.

10. Menani JV, Saad WA, Camargo LAA, Antunes-Rodrigues J, Covian MR & Saad WA (1984). Effect of cholinergic and adrenergic stimulation of the subfornical organ on water intake. Pharmacology, Biochemistry and Behavior, 20: 301-306.

11. Fregly MJ, Rowland NE & Cade JR (1992). Comparison of the hyperhydrating effects of angiotensin II and isoproterenol. Pharmacology, Biochemistry and Behavior, 43: 1143-1149.

12. Antunes-Rodrigues J & McCann SM (1970). Water, sodium and food intake induced by injections of cholinergic and adrenergic drugs into the third ventricle of rat brain. Proceedings of the Society for Experimental Biology and Medicine, 133: 1464-1470.

13. Bielarczyk H, Tomsig JL & Suszkiw JB (1994). Perinatal low level lead exposure and the septal-hippocampal cholinergic system: selective reduction of muscarinic receptors and cholineacetyltransferase. Brain Research, 643: 211-217.

14. Lasley SM & Lane JD (1988). Diminished regulation of mesolimbic dopaminergic activity in rat after chronic inorganic lead exposure. Toxicology and Applied Pharmacology, 95: 474-483.

15. Alkondon M, Costa ACS, Radhakrishnan V, Aronstam RS & Albuquerque EX (1990). Selective blockade of NMDA-activated currents may be implicated in learning deficits caused by lead. FEBS Letters, 261: 124-130.

16. Kumar MV & Desiraju T (1990). Regional alterations of brain biogenic amines and GABA/glutamate levels following chronic lead exposure during neonatal development. Archives of Toxicology, 64: 305-314.

17. Collins MF, Hrdina PD, Whittle E & Singhal RL (1984). The effects of low-level lead exposure in developing rats: change in circadian locomotor activity and hippocampal noradrenaline turnover. Canadian Journal of Physiology and Pharmacology, 62: 430-435.

18. Bellin SI, Bhatnagar RK & Johnson AK (1987). Periventricular noradrenergic systems are critical for angiotensin-induced drinking and blood pressure responses. Brain Research, 403: 105-112.

19. Bellin SI, Landas SK & Johnson AK (1988). Selective catecholamine depletion of structures along the ventral lamina terminalis: effects on experimentally-induced drinking and pressor responses. Brain Research, 456: 9-16.

Acknowledgments

We are indebted to Mr. Vanilson Souza for technical assistance and to Mr. José de Souza for animal care.

Address for correspondence: J.B. Fregoneze, Departamento de Zoologia, Instituto de Biologia, Universidade Federal da Bahia, 40170-110 Salvador, BA, Brasil.

Presented at the XI Annual Meeting of the Federação de Sociedades de Biologia Experimental, Caxambu, MG, Brasil, August 21-24, 1996. Research supported by CNPq (Nos. 300772/86-6 and 301099/92-8). Received April 11, 1996. Accepted December 17, 1996.

• 2. Bradbury MWB & Deane R (1993). Permeability of the blood-brain barrier to lead. Neurotoxicology, 14: 131-136.
• 3. Arvidson B (1994). A review of axonal transport of metals. Toxicology, 88: 1-14.
• 4. Luthman J, Lidqvist E, Gerhardt GA, Olson L & Hoffer BH (1994). Alterations in central monoamine systems after postnatal lead acetate treatment in rats. Environmental Research, 65: 100-118.
• 5. Sutoo D, Akiyama K & Maeda I (1987). The effect of cadmium administration on the biogenic amine distributions and amounts in the mouse brain. Kitasato Archives of Experimental Medicine, 60: 157-166.
• 6. Rastogi RB, Merali Z & Singhal RI (1977). Cadmium alters behaviour and the biosynthetic capacity for catecholamines and serotonin in neonatal rat brain. Journal of Neurochemistry, 28: 789-794.
• 7. Fregoneze JB, Cunha M, Bulcăo C, Ferreira H & De Castro e Silva E (1994). Acute effect of intracerebroventricular administration of lead on the drinking behavior of rats induced by dehydration or central cholinergic and angiotensinergic stimulation. Physiology and Behavior, 56: 129-133.
• 8. De Castro e Silva E, Ferreira H, Cunha M, Bulcăo C, Sarmento C, De Oliveira IR & Fregoneze JB (1996). Effect of central acute administration of cadmium on drinking behavior. Pharmacology, Biochemistry and Behavior, 53: 687-693.
• 9. Callera JC, Saad WA, Camargo LAA, Renzi A, De Luca Jr LA & Menani JV (1994). Role of adrenergic pathways of the lateral hypothalamus on water intake and pressor response induced by the cholinergic activation of the medial septal area in rats. Neuroscience Letters, 167: 153-155.
• 10. Menani JV, Saad WA, Camargo LAA, Antunes-Rodrigues J, Covian MR & Saad WA (1984). Effect of cholinergic and adrenergic stimulation of the subfornical organ on water intake. Pharmacology, Biochemistry and Behavior, 20: 301-306.
• 11. Fregly MJ, Rowland NE & Cade JR (1992). Comparison of the hyperhydrating effects of angiotensin II and isoproterenol. Pharmacology, Biochemistry and Behavior, 43: 1143-1149.
• 12. Antunes-Rodrigues J & McCann SM (1970). Water, sodium and food intake induced by injections of cholinergic and adrenergic drugs into the third ventricle of rat brain. Proceedings of the Society for Experimental Biology and Medicine, 133: 1464-1470.
• 13. Bielarczyk H, Tomsig JL & Suszkiw JB (1994). Perinatal low level lead exposure and the septal-hippocampal cholinergic system: selective reduction of muscarinic receptors and cholineacetyltransferase. Brain Research, 643: 211-217.
• 14. Lasley SM & Lane JD (1988). Diminished regulation of mesolimbic dopaminergic activity in rat after chronic inorganic lead exposure. Toxicology and Applied Pharmacology, 95: 474-483.
• 15. Alkondon M, Costa ACS, Radhakrishnan V, Aronstam RS & Albuquerque EX (1990). Selective blockade of NMDA-activated currents may be implicated in learning deficits caused by lead. FEBS Letters, 261: 124-130.
• 16. Kumar MV & Desiraju T (1990). Regional alterations of brain biogenic amines and GABA/glutamate levels following chronic lead exposure during neonatal development. Archives of Toxicology, 64: 305-314.
• 17. Collins MF, Hrdina PD, Whittle E & Singhal RL (1984). The effects of low-level lead exposure in developing rats: change in circadian locomotor activity and hippocampal noradrenaline turnover. Canadian Journal of Physiology and Pharmacology, 62: 430-435.
• 18. Bellin SI, Bhatnagar RK & Johnson AK (1987). Periventricular noradrenergic systems are critical for angiotensin-induced drinking and blood pressure responses. Brain Research, 403: 105-112.
• 19. Bellin SI, Landas SK & Johnson AK (1988). Selective catecholamine depletion of structures along the ventral lamina terminalis: effects on experimentally-induced drinking and pressor responses. Brain Research, 456: 9-16.

Correspondence and Footnotes

Publication Dates

• Publication in this collection
  19 Mar 1999
• Date of issue
  Mar 1997

History

• Accepted
  17 Dec 1996
• Received
  11 Apr 1996