Potential Applications of Neuroendocrine ... - ACS Publications

Chapter 11

Potential Applications of Neuroendocrine Research to Insect Control

Downloaded by CORNELL UNIV on September 3, 2016 | http://pubs.acs.org Publication Date: November 22, 1988 | doi: 10.1021/bk-1988-0379.ch011

Larry L. Keeley Laboratories for Invertebrate Neuroendocrine Research, Department of Entomology, Texas Agricultural Experiment Station, Texas A&M University, College Station, TX 77843 Advances in biotechnology that result from neuroendocrine research may have applications for the control of insect pests. Neurohormones are the master regulatory hormones of insects and affect c r i t i c a l physiological processes that include: molting, metamorphosis, reproduction, and general homeostasis. Manipulation of neurohormone titers would disrupt sensitive physiological processes and kill or debilitate treated insects. Sensitive neuroendocrine events include: hormone synthesis, hormone secretion and degradation; hormone-target cell-receptor interactions and target cell responses. Neuroendocrine events and titers may be altered by: (1) genetic engineering of neurohormone genes into suitable baculovirus cloning-expression vectors; (2) disruption of neurosecretory activity by exogenous chemicals that affect aminergic neurons; (3) development of analogs that are antagonists or superagonists of neurohormones at receptor proteins; and (4) inhibition of enzymes instrumental in hormone synthesis or degradation.

The r o l e o f the a n i m a l b r a i n as a s o u r c e f o r e n d o c r i n e f a c t o r s was d i s c o v e r e d i n i n s e c t s w i t h t h e p i o n e e r i n g work o f Kopec i n 1917 {V). He found t h a t l i g a t i o n o f t h e head p r e v e n t e d m o l t i n g and metamorphosis i n gypsy moth l a r v a e . I n 1941, t h e phenomenon o f n e u r o s e c r e t i o n was d e s c r i b e d f o r i n s e c t s b y B e r t a S c h a r r e r (2^) u s i n g the c o c k r o a c h , Leucophaea maderae. D e s p i t e t h i s e a r l y use o f i n s e c t s as models f o r s t u d y i n g n e u r o e n d o c r i n e p r o c e s s e s , t h e t o p i c of i n s e c t neuroendocrinology has lagged b e h i n d i t s v e r t e b r a t e c o u n t e r p a r t because o f t h e s c a r c i t y o f i d e n t i f i e d neurohormones. Over the f o u r decades s i n c e S c h a r r e r f i r s t d e s c r i b e d t h e n e u r o s e c r e t o r y r o l e f o r c o c k r o a c h b r a i n n e u r o n s , about two dozen p h y s i o l o g i c a l p r o c e s s e s o f i n s e c t s have been i d e n t i f i e d as b e i n g n e u r o e n d o c r i n e dependent (3). However, i t was n o t u n t i l 1975-76 t h a t t h e f i r s t i n s e c t neurohormone s t r u c t u r e s were d e f i n e d (4,5)· No f u r t h e r s t r u c t u r e s were r e p o r t e d u n t i l 1984 ( 6 - 8 ) . At present,

0097-6156/88/Ό379-0147$06.00/0 « 19.88 American Chemical Society

American Chemical Society Library 15th St., for N.W. Hedin et 1155 al.; Biotechnology Crop Protection ACS Symposium Series; American Chemical Society: Washington, DC, 1988. Washington, D.C. 20036


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t h e r e are about two dozen neurohormonal s t r u c t u r e s r e p o r t e d i n the l i t e r a t u r e , b u t many o f t h e s e are n a t u r a l analogs (bioanalogs) of each o t h e r and a f f e c t the same p h y s i o l o g i c a l p r o c e s s e s . Neurohormones a r e the master r e g u l a t o r s f o r p h y s i o l o g i c a l p r o c e s s e s i n b o t h v e r t e b r a t e and i n v e r t e b r a t e a n i m a l s . In i n s e c t s , neurohormones r e g u l a t e g e n e r a l h o m e o s t a t i c a c t i v i t i e s such a s : b e h a v i o r ; the s y n t h e s i s o f c i r c u l a t i n g c a r b o h y d r a t e s , l i p i d s and p r o t e i n s ; water and s a l t b a l a n c e ; h e a r t b e a t r a t e ; i n v o l u n t a r y muscle c o n t r a c t i o n s and b a s a l m e t a b o l i s m . Neurohormones a l s o r e g u l a t e the s y n t h e s i s and s e c r e t i o n o f the e c d y s t e r o i d s and j u v e n i l e hormones t h a t d i r e c t l y c o n t r o l the p r o c e s s e s o f m o l t i n g , metamorphosis and reproduction. F u r t h e r m o r e , neurohormones a f f e c t s p e c i f i c e v e n t s o f growth and r e p r o d u c t i o n such as e c l o s i o n , c u t i c l e t a n n i n g , o v u l a t i o n and o v i p o s i t i o n . The p o t e n t i a l f o r u s i n g e n d o c r i n e imbalance as a means o f i n s e c t p e s t c o n t r o l was s u g g e s t e d by W i l l i a m s (9,10)· He p r o p o s e d t h a t e x p o s i n g immature i n s e c t s to j u v e n i l e hormone (JH) a t the time o f metamorphosis, when JH i s n o r m a l l y a b s e n t , would cause abnormal development and i n d i v i d u a l s i n c a p a b l e o f s u r v i v a l . S i n c e i n s e c t metamorphosis i s u n i q u e , JH d i s r u p t i o n would a f f e c t o n l y i n s e c t s . T h i s would r e s u l t i n an e n v i r o n m e n t a l l y s a f e a p p r o a c h t o i n s e c t c o n t r o l as compared to c u r r e n t c h e m i c a l p e s t i c i d e s which are l e s s i n s e c t s p e c i f i c and more b i o c i d a l . The JH approach t o p e s t i n s e c t c o n t r o l i s most e f f e c t i v e when a d u l t s are the d e s t r u c t i v e s t a g e , and commercial p r e p a r a t i o n s o f JH mimics are a v a i l a b l e f o r use i n the c o n t r o l o f a d u l t f l i e s , m o s q u i t o e s , and f l e a s and, r e c e n t l y , f o r cockroach reproduction. However, many p e s t i n s e c t s a r e d e s t r u c t i v e as l a r v a e . The use o f neurohormones f o r p e s t c o n t r o l remains s p e c u l a t i v e , b u t c o n s i d e r a b l e i n t e r e s t i s d e v e l o p i n g c o n c e r n i n g the p o s s i b i l i t y , and s e v e r a l a r t i c l e s have p r o p o s e d approaches f o r a p p l y i n g n e u r o e n d o c r i n e i n f o r m a t i o n (11-14)· Examples e x i s t i n the l i t e r a t u r e which s u g g e s t t h a t n e u r o e n d o c r i n e imbalance a f f e c t s insects adversely. M o l t i n g c e a s e s a f t e r d e s t r u c t i o n o f the n e u r o s e c r e t o r y c e l l s t h a t s e c r e t e the p r o t h o r a c i c o t r o p i c hormone (PTTH) (_1_f0 , and removal o f the c o r p o r a c a r d i a c a as a n e u r o e n d o c r i n e s o u r c e a f f e c t s the p h y s i o l o g i c a l performance o f a c t i v e i n s e c t s . For example, the c o r p o r a c a r d i a c a c o n t a i n a neurohormone t h a t m o b i l i z e s t r e h a l o s e , the major i n s e c t b l o o d sugar (16,17). Normal C a l l i p h o r a e r y t h r o c e p h a l a b l o w f l i e s f l y f o r more than 45 min w i t h o u t e x h a u s t i o n o r a measurable d e c l i n e i n hemolymph t r e h a l o s e , b u t cardiacectomized b l o w f l i e s have an 85% r e d u c t i o n i n t r e h a l o s e and a r e exhausted a f t e r 45 min o f f l i g h t (J_8 ). s i m i l a r l y , removal o f the c o r p o r a c a r d i a c a r e d u c e s f l i g h t speed by 40% i n l o c u s t s (J_9), p r o b a b l y because o f a d e f i c i e n c y i n the a d i p o k i n e t i c hormone (ΑΚΗ) t h a t m o b i l i z e s f a t body l i p i d s as an energy s o u r c e f o r the f l i g h t m u s c l e s . A l t e r n a t i v e l y , e x c e s s i v e amounts o f neurohormones appear d e b i l i t a t i n g , as w e l l . L a r g e doses o f p r o c t o l i n cause temporary s t u p o r and i m m o b i l i t y i n c o c k r o a c h e s (^0_). I t i s a g r e a t l e a p from o b s e r v i n g e x p e r i m e n t a l r e s p o n s e s t o e n d o c r i n e imbalance i n the l a b o r a t o r y t o c o n t r o l l i n g i n s e c t s i n the f i e l d by n e u r o e n d o c r i n e d i s r u p t i o n . N e v e r t h e l e s s , laboratory e x p e r i m e n t s i n d i c a t e t h a t s u s t a i n e d n e u r o e n d o c r i n e d i s r u p t i o n would have p r o f o u n d e f f e c t s on growth, development and p h y s i o l o g i c a l functions. Whereas expérimenta! p r o c e d u r e s such as i n j e c t i o n o f a

Hedin et al.; Biotechnology for Crop Protection ACS Symposium Series; American Chemical Society: Washington, DC, 1988.

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n a t u r a l hormone do not e x h i b i t extreme r e s p o n s e s because the hormone i s degraded by p r o t e a s e s , a g e n t s d e s i g n e d t o d i s r u p t the n e u r o e n d o c r i n e b a l a n c e f o r c o n t r o l purposes w i l l have to be s t a b l e i n b o t h the environment and the i n s e c t . Such a g e n t s w i l l cause a p e r s i s t e n t e n d o c r i n e imbalance t h a t r e s u l t s i n the t a r g e t t i s s u e receiving e i t h e r a sustained, uncontrolled hyperstimulation or suppression of response. T h i s p e r s i s t e n t , u n c o n t r o l l e d response by the t a r g e t t i s s u e w i l l cause s e r i o u s d e b i l i t a t i o n and, u l t i m a t e l y , d e a t h o f the t r e a t e d i n s e c t .


Objections

to U s i n g N e u r o e n d o c r i n e - B a s e d P e s t I n s e c t

Control

P e p t i d e N a t u r e of the Neurohormones. Neurohormones are p r o t e i n s and, as such, are i n t r i n s i c a l l y u n s t a b l e and u n s u i t e d f o r a p p l i c a t i o n i n the environment or d i r e c t l y t o i n s e c t s f o r p e s t c o n t r o l purposes. P e p t i d e s are e a s i l y degraded i n the environment by l i g h t , h e a t , and m i c r o o r g a n i s m s . A l s o , i t i s u n l i k e l y t h a t n a t u r a l neurohormones c o u l d g a i n e n t r y i n t o exposed i n s e c t s . Most p r o t e i n s c a n n o t p e n e t r a t e the i n s e c t c u t i c l e , and t h e y would be d i g e s t e d t o t h e i r c o n s t i t u e n t amino a c i d s by gut p r o t e a s e s i f consumed o r a l l y . For example, t o p i c a l l y a p p l i e d p r o c t o l i n i s n o t a b s o r b e d by M, s e x t a l a r v a e and i n g e s t e d p r o c t o l i n i s degraded r a p i d l y ( 2J_). Therefore, i t i s highly u n l i k e l y that n a t u r a l neurohormones c o u l d be used t o a f f e c t the n e u r o e n d o c r i n e b a l a n c e o f target insects. However, i t may be p o s s i b l e t o overcome some o f the disadvantages of peptides. P e p t i d e a n a l o g s might be s t a b i l i z e d so as to r e s i s t d i g e s t i v e p r o t e a s e s and be permeable t o the i n s e c t g u t . A n a l o g s might be d e s i g n e d t o be s t a b l e o r permeable t o the c u t i c l e i n the same manner as p r e s e n t i n s e c t i c i d e s . An example e x i s t s o f a s e r i e s o f p e p t i d e s t h a t were s y n t h e s i z e d w i t h i s o p r e n e - o r t e r p e n o i d - l i k e s t r u c t u r e s and e x h i b i t e d j u v e n i l e hormone a c t i v i t y when a p p l i e d t o p i c a l l y t o l a r v a l and p u p a l i n s e c t s (22). Finally, the use of i n s e c t v i r u s e s as h i g h l y - e f f i c i e n t c l o n i n g - e x p r e s s i o n v e c t o r s f o r neurohormone genes might p r o v i d e a v e h i c l e f o r c a r r y i n g the genes d i r e c t l y i n t o the i n s e c t where the hormone c o u l d be p r o d u c e d i n u n c o n t r o l l e d superabundance. U n i v e r s a l i t y o f Neurohormones Among A n i m a l s . I f b o t h i n s e c t s and v e r t e b r a t e s share common p e p t i d e s t r u c t u r e s as neurohormones, then s t r a t e g i e s f o r m a n i p u l a t i n g i n s e c t neurohormones c o u l d be d e t r i m e n t a l to n o n t a r g e t a n i m a l s and p r e s e n t s e r i o u s e n v i r o n m e n t a l concerns. However, p r e s e n t f i n d i n g s do not s u p p o r t the c o n t e n t i o n t h a t i n s e c t s and v e r t e b r a t e s s h a r e common hormones f o r the same functions. A n t i b o d i e s t o v e r t e b r a t e p e p t i d e hormones r e a c t w i t h p e p t i d e s p r e s e n t i n i n s e c t s {23)· But, i n those c a s e s where an immunoreactive v e r t e b r a t e - l i k e p e p t i d e has been o b t a i n e d , the p e p t i d e does not have the p h y s i o l o g i c a l a c t i o n found i n i t s vertebrate counterpart. C o n v e r s e l y , i n those c a s e s where a p h y s i o l o g i c a l p r o c e s s was used as the b a s i s f o r i s o l a t i o n o f an i n s e c t neurohormone, the r e s u l t i n g p e p t i d e has had a unique structure. S i m i l a r i t i e s a r e sometimes noted between the p e p t i d e s t r u c t u r e s o f i n s e c t and v e r t e b r a t e neurohormones. For example, l e u c o s u l f a k i n i n i s an i n s e c t m y o t r o p i n t h a t has a 50% i d e n t i t y w i t h v e r t e b r a t e g a s t r i n - c h o l e c y s t o k i n i n (2A)· I n t e s t i n a l f u n c t i o n s are


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s t i m u l a t e d i n v e r t e b r a t e s by g a s t r i n and c h o l e c y s t o k i n i n and i n i n s e c t s by l e u c o s u l f a k i n i n ; however, n e i t h e r g a s t r i n nor c h o l e c y s t o k i n i n a f f e c t i n t e s t i n a l c o n t r a c t i o n s i n the i n s e c t - g u t bioassay. The 4 kDa PTTH from s i l k w o r m s a l s o has an a p p r o x i m a t e l y 50% i d e n t i t y w i t h v e r t e b r a t e i n s u l i n (25), y e t v e r t e b r a t e i n s u l i n does not a c t i v a t e the p r o t h o r a c i c g l a n d s a t d o s e s t h a t exceed normal PTTH t i t e r s by 10,000 t i m e s . V e r t e b r a t e - l i k e n e u r o p e p t i d e s are p r e s e n t i n i n s e c t s as d e m o n s t r a t e d i m m u n o l o g i c a l l y , and p r o b a b l y v i c e v e r s a , b u t i t i s u n c l e a r what the f u n c t i o n i s f o r t h e s e p e p t i d e s i n t h e i r h e t e r o l o g o u s a n i m a l s y s t e m . The s t r u c t u r a l s i m i l a r i t i e s between t h e s e m o l e c u l e s s u g g e s t t h a t common, b i o l o g i c a l l y - a c t i v e a n c e s t r a l m o l e c u l e s may have e x i s t e d and e v o l v e d t o p e r f o r m d i f f e r e n t f u n c t i o n s d e p e n d i n g on the p h y s i o l o g i c a l d i v e r s i t y and needs o f the animals i n v o l v e d . I t i s s t i l l p o s s i b l e t o make use o f the n e u r o e n d o c r i n e a p p r o a c h even i f the p r o c e d u r e s used a f f e c t b o t h i n s e c t and v e r t e b r a t e n e u r o e n d o c r i n e systems. F o r example, most c u r r e n t i n s e c t i c i d e s a r e g e n e r a l n e r v e p o i s o n s o f h i g h e r a n i m a l s . The broad-based t o x i c i t y o f t h e s e c h e m i c a l s has been m i n i m i z e d by d i r e c t i n g the i n s e c t i c i d e s a t t a r g e t organisms t h r o u g h s p e c i f i c a p p l i c a t i o n and formulation procedures. Such d i r e c t e d a p p l i c a t i o n p r o c e d u r e s w i l l s u r e l y be p a r t o f the s t r a t e g i e s d e v e l o p e d f o r d i s r u p t i n g n e u r o e n d o c r i n e processes i n i n s e c t s . Neurohormone D i s t r i b u t i o n Among I n s e c t S p e c i e s . I t i s important to c o n s i d e r the d i s t r i b u t i o n o f the v a r i o u s neurohormones among the i n s e c t s p e c i e s when d e v e l o p i n g a n e u r o e n d o c r i n e - b a s e d c o n t r o l strategy. Some hormones such as the n e u r o p e p t i d e s t h a t r e g u l a t e m o l t i n g must be u n i v e r s a l s i n c e a l l i n s e c t s grow by m o l t i n g . On the o t h e r hand, some hormones may be r e s t r i c t e d i n t h e i r d i s t r i b u t i o n . I t i s l i k e l y t h a t a q u a t i c i n s e c t s and s t o r e d g r a i n i n s e c t s may have l i t t l e need f o r a n t i d i u r e t i c and d i u r e t i c f a c t o r s , r e s p e c t i v e l y . In a d d i t i o n , hormones w i t h the same f u n c t i o n may d i f f e r s t r u c t u r a l l y between s p e c i e s , and f a m i l i e s o f hormones e x i s t . The adipokinetic/hypertrehalosemic (AKH/HTH) hormones c o n s t i t u t e a p e p t i d e f a m i l y t h a t , now, has e i g h t r e p r e s e n t a t i v e s t r u c t u r e s from e i g h t s p e c i e s o f i n s e c t s . One o t h e r ΑΚΗ-related s t r u c t u r e ( r e d p i g m e n t - c o n c e n t r a t i n g hormone) i s found i n c r u s t a c e a n s and c r o s s r e a c t s i n i n s e c t s . In some c a s e s , the same hormone i s found i n two d i s t i n c t i n s e c t s p e c i e s ; i n o t h e r c a s e s two AKH/HTH p e p t i d e s w i t h d i f f e r e n t s t r u c t u r e s and f u n c t i o n s a r e found i n the same species. In a l l c a s e s , the AKH/HTH b i o a n a l o g s a f f e c t l i p i d o r c a r b o h y d r a t e m e t a b o l i s m by the f a t body, a l t h o u g h the d i f f e r e n t hormones have d i f f e r i n g p o t e n c i e s among the v a r i o u s s p e c i e s f o r t h e i r metabolic e f f e c t s . Advantages t o U s i n g N e u r o e n d o c r i n e - B a s e d P e s t

Insect

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There are a number of advantages t o a n e u r o e n d o c r i n e approach to i n s e c t p e s t management. 1. Neurohormones r e g u l a t e numerous c r i t i c a l p h y s i o l o g i c a l p r o c e s s e s s u s c e p t i b l e to a l t e r a t i o n .


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2. Neurohormones a r e amenable t o g e n e t i c e n g i n e e r i n g t e c h n o l o g y because o f t h e i r p e p t i d i c n a t u r e . 3. S e v e r a l n e u r o e n d o c r i n e e v e n t s a r e s u s c e p t i b l e t o a l t e r a t i o n . 4. Neurohormones a r e s t r u c t u r a l l y d i v e r s e so t h a t i t i s p o s s i b l e t o s y n t h e s i z e a wide a r r a y o f a g o n i s t i c o r a n t a g o n i s t i c a n a l o g s . Endocrine Regulated P h y s i o l o g i c a l Processes. The v a r i e t y o f p h y s i o l o g i c a l p r o c e s s e s a f f e c t e d by neurohormones s u g g e s t s t h a t d i s r u p t i o n s i n n e u r o e n d o c r i n e - d e p e n d e n t p r o c e s s e s may be used as the = b a s i s f o r a new a p p r o a c h t o i n s e c t p e s t c o n t r o l . S i n c e neurohormones such as PTTH and a l l a t o t r o p i n and a l l a t o s t a t i n r e g u l a t e the s e c r e t i o n o f t h e e c d y s t e r o i d s and JH, m a n i p u l a t i o n o f t h e s e neurohormones c o u l d i n f l u e n c e m o l t i n g and metamorphosis i n l a r v a e and r e p r o d u c t i o n i n a d u l t s j u s t as would m a n i p u l a t i o n o f the e c d y s t e r o i d s o r the J H s , themselves. D i s r u p t i o n o f e c l o s i o n hormone c o u l d p r e v e n t t h e o n s e t o f m o l t i n g b e h a v i o r and e c d y s i s , and d i s r u p t i o n o f b u r s i c o n c o u l d p r e v e n t t a n n i n g o f the s o f t c u t i c l e o f newly-molted i n d i v i d u a l s . D i s r u p t i o n o f e i t h e r e c d y s i s o r t a n n i n g would r e s u l t i n the d e a t h o f affected insects. Enhancement o f d i u r e t i c hormone a c t i o n s c o u l d be f a t a l f o r those i n s e c t s t h a t have c r i t i c a l problems o f m a i n t a i n i n g body w a t e r , esp. soft-bodied l a r v a e . S o f t - b o d i e d l a r v a e r e l y on hemolymph volume t o m a i n t a i n t u r g o r f o r movement, and d e h y d r a t i o n would r e s u l t i n i m m o b i l i t y f o l l o w e d by d e a t h . I n h i b i t i o n of a n t i d i u r e t i c hormones c o u l d have s e r i o u s e f f e c t s on i n s e c t s t h a t r e s i d e i n e x t r e m e l y d r y e n v i r o n m e n t s , e.g. s t o r e d g r a i n . A n t i d i u r e t i c hormone a n t a g o n i s t s i n c o m b i n a t i o n w i t h d i u r e t i c hormone a g o n i s t s would be e s p e c i a l l y potent. D i s r u p t i o n o f n e u r o p e p t i d e s t h a t a f f e c t muscle c o n t r a c t i o n might r e s u l t i n i m m o b i l i t y o r b l o c k i n t e s t i n a l p e r i s t a l s i s t o i n h i b i t f e e d i n g and d i g e s t i o n . P r o c t o l i n - i n d u c e d i m m o b i l i t y i n c o c k r o a c h e s i l l u s t r a t e s t h i s s i t u a t i o n (^0)· D i s r u p t i o n o f ΑΚΗ o r HTH c o u l d p r e v e n t the r e l e a s e o f l i p i d s and c a r b o h y d r a t e s from t h e f a t body and t h e i r use as e n e r g y f u e l s for mobility. F l i g h t speed, wingbeat f r e q u e n c y and endurance d e c r e a s e d i n i n s e c t s r e n d e r e d d e f i c i e n t i n ΑΚΗ o r HTH by c o r p o r a c a r d i a c a removal (18,19)· A l t h o u g h l o w e r i n g b l o o d m e t a b o l i t e s might have a r e l a t i v e l y b e n i g n e f f e c t on s u r v i v a l , t h e r e s u l t i n g d e c r e a s e i n m o b i l i t y would make t r e a t e d i n s e c t s more s u s c e p t i b l e t o e n v i r o n m e n t a l m o r t a l i t y f a c t o r s , such as p a r a s i t i z a t i o n , prédation and a d v e r s e weather and p r e v e n t m i g r a t i o n o r movement f o r l o n g d i s t a n c e s i n s e a r c h o f f o o d , s h e l t e r , o r mates. S e v e r a l s t e p s i n the r e p r o d u c t i v e p r o c e s s a r e neurohormone regulated. Pheromone s y n t h e s i s - r e l e a s e , o v u l a t i o n , o v i p o s i t i o n , and gonad m a t u r a t i o n might a l s o be c o n s i d e r e d as n e u r o e n d o c r i n e dependent p r o c e s s e s s u s c e p t i b l e t o m a n i p u l a t i o n f o r r e p r o d u c t i v e disruption. Recent s t u d i e s have d e m o n s t r a t e d t h a t a g i v e n hormone may have m u l t i p l e e f f e c t s . F o r example, f o r over 20 y e a r s we have known t h a t f a c t o r s e x i s t e d i n the c o r p o r a c a r d i a c a o f c o c k r o a c h e s t h a t a f f e c t e d the h e a r t b e a t r a t e and t r e h a l o s e l e v e l s o f the hemolymph. O n l y w i t h the i s o l a t i o n - c h a r a c t e r i z a t i o n o f the two AKH/HTH f a c t o r s from the c o r p o r a c a r d i a c a o f ]?. americana d i d we f i n d t h a t the h e a r t b e a t and c a r b o h y d r a t e r e g u l a t i o n s a r e performed by i d e n t i c a l f a c t o r s (6-8)· F i n a l l y , we have found i n my l a b o r a t o r y t h a t the HTH o f the


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c o c k r o a c h B l a b e r u s d i s c o i d a l i s a f f e c t s not o n l y t r e h a l o s e and h e a r t b e a t r a t e but a l s o heme s y n t h e s i s f o r cytochrome i n the f a t body ( u n p u b l i s h e d data).

synthesis production

Endocrine Events S u s c e p t i b l e to Manipulation. There a r e a s e r i e s o f e v e n t s which are common to the p r o d u c t i o n and a c t i o n o f any hormone. These e v e n t s i n c l u d e : hormone s y n t h e s i s , s e c r e t i o n and t r a n s p o r t ; h o r m o n e - t a r g e t c e l l r e c e p t o r i n t e r a c t i o n ; t a r g e t c e l l r e s p o n s e ; and hormone d e g r a d a t i o n . S e v e r a l of these events are s u s c e p t i b l e to m a n i p u l a t i o n by e x t e r n a l i n f l u e n c e s t h a t might be d e v i s e d i n t o control strategies. Neurohormones are p r o t e i n s and a r e , t h e r e f o r e , u l t i m a t e l y gene p r o d u c t s . I t seems q u e s t i o n a b l e t h a t the t r a n s c r i p t i o n a l l a c t i v i t y o f neurohormone genes c o u l d be d i s r u p t e d by a means t h a t would be s p e c i f i c t o i n s e c t s and s a f e f o r g e n e r a l u s e . However, i f we were t o f i n d t h a t the neurohormone genes are r e g u l a t e d by i n s e c t hormones such as the e c d y s t e r o i d s o r JHs, then i t might be p o s s i b l e t o s u p p r e s s i n s e c t neurohormone gene a c t i v i t y s e l e c t i v e l y . The enzymes t h a t p r o c e s s prohormones i n t o a c t i v e hormones are candidates for a r t i f i c i a l manipulation. N e u r o e n d o c r i n e r e s e a r c h on v e r t e b r a t e s and n o n i n s e c t i n v e r t e b r a t e s i n d i c a t e s t h a t the p r i m a r y t r a n s l a t i o n p r o d u c t s from i n s e c t neurohormone genes can be e x p e c t e d t o be l a r g e prohormones t h a t a r e p o s t - t r a n s l a t i o n a l l y p r o c e s s e d i n t o the a c t i v e hormone by cleavage a t p a i r s of b a s i c amino a c i d s . No e v i d e n c e i s p u b l i s h e d on the n a t u r e o f the i n s e c t prohormones, a l t h o u g h s e v e r a l l a b o r a t o r i e s a r e examining t h i s t o p i c . N o t h i n g i s known about the prohormone p r o c e s s i n g enzymes o f i n s e c t s b u t hormone p r o c e s s i n g enzymes a r e r e p o r t e d f o r v e r t e b r a t e s (26,27), and i t i s p r o b a b l e t h a t the enzymes are s i m i l a r f o r the two g r o u p s . A g a i n , i f the p r o c e s s i n g enzymes are s i m i l a r , then t h e s e enzymes may not be the i d e a l s i t e f o r d i r e c t i n g a c o n t r o l p r o c e d u r e s i n c e such an a p p r o a c h might a l s o a f f e c t n o n t a r g e t o r g a n i s m s . N e u r o s e c r e t i o n i s s u s c e p t i b l e t o m a n i p u l a t i o n t o a l t e r the neuroendocrine balance. Neurohormones a r e s e c r e t e d from t h e i r s o u r c e neurons under the d i r e c t i o n o f o t h e r e n d o c r i n e a g e n t s o r i n r e s p o n s e to nervous i n p u t from r e g u l a t o r y n e u r o n s . The r e g u l a t o r y neurons a f f e c t n e u r o s e c r e t o r y a c t i v i t y v i a s p e c i f i c b i o g e n i c amine neurotransmitters. V e r t e b r a t e s t u d i e s show t h a t b i o g e n i c amine n e u r o t r a n s m i t t e r s s t i m u l a t e s p e c i f i c neurohormone s e c r e t i o n (28) P e p t i d i c r e l e a s i n g f a c t o r s a l s o promote s p e c i f i c n e u r o s e c r e t i o n (29) i n v e r t e b r a t e s , b u t such r e l e a s i n g f a c t o r s a r e unknown i n i n s e c t s . D i s r u p t i o n o f neurohormone s e c r e t i o n p a t t e r n s i n i n s e c t s , i f performed a t c r i t i c a l times i n the l i f e c y c l e , c o u l d d i s t u r b s e n s i t i v e p h y s i o l o g i c a l e v e n t s such as m o l t i n g , metamorphosis or reproduction. L i t t l e i s known about the t r a n s p o r t o f i n s e c t neurohormones from t h e i r s o u r c e t o t a r g e t t i s s u e s . Presumably, the hormones a r e t r a n s p o r t e d v i a the hemolymph, a l t h o u g h t h e r e a r e i n d i c a t i o n s o f t r a n s p o r t by n e r v e s e x t e n d i n g from the e n d o c r i n e s o u r c e t o the t a r g e t t i s s u e ( 30_) o r by hemocytes ( 3J_). Hemolymph-borne hormones s h o u l d be s u s c e p t i b l e t o d e g r a d a t i o n by c i r c u l a t i n g p r o t e a s e s , b u t t h i s may not be the c a s e . The hormones may be bound t o a p r o t e c t i n g c a r r i e r p r o t e i n ( n e u r o p h y s i n ) , or s p e c i f i c d e g r a d a t i v e p r o t e a s e s may be a s s o c i a t e d w i t h r e c e p t o r s i t e s on t a r g e t c e l l membranes so t h a t the c i r c u l a t i n g hormone i s n o t a f f e c t e d . The p r e s e n c e o f the amide



11.

KEELEY


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and p y r o - g l u t a m a t e groups a t the C- and N - t e r m i n i o f the AKH/HTH hormones may s e r v e a p r o t e c t i v e r o l e t o p r e v e n t d e g r a d a t i o n d u r i n g transport. S i n c e neurohormones a r e p r o t e i n a c e o u s , i t i s u n l i k e l y t h a t t h e y can be a p p l i e d d i r e c t l y t o an i n s e c t w i t h the hope o f i m p l e m e n t i n g an e n d o c r i n e imbalance; however, i t i s f e a s i b l e t o d e s i g n and d e v e l o p s t a b l e , n o n p e p t i d e a n a l o g s t o the a c t i v e c o n f o r m a t i o n o f the hormone. The use o f s t a b l e a n a l o g s t h a t e i t h e r mimic o r b l o c k the neurohormone r e c e p t o r s i t e s o f t a r g e t t i s s u e s c o n s t i t u t e one o f the most p r o m i s i n g means f o r m a n i p u l a t i n g e n d o c r i n e - d e p e n d e n t p r o c e s s e s i n an i n s e c t c o n t r o l s t r a t e g y . A l t e r n a t i v e l y , i n h i b i t i o n o f the a c t i v e s i t e o f d e g r a d a t i v e p r o t e a s e enzymes by neurohormone a n a l o g s c o u l d r e s u l t i n the a c c u m u l a t i o n o f n a t u r a l hormones and the hyperstimulation of sensitive processes. Neuroendocrine

Manipulation

Four ways t h a t neurohormones and neurohormone-dependent p h y s i o l o g i c a l p r o c e s s e s m i g h t be a f f e c t e d a r e : 1. g e n e t i c e n g i n e e r i n g and recombinant DNA b i o t e c h n o l o g y ; 2. a l t e r e d n e u r o s e c r e t i o n ; 3. i n h i b i t i o n o f enzymes f o r hormone s y n t h e s i s o r d e g r a d a t i o n ; and 4. neurohormone a n a l o g s . Recombinant DNA B i o t e c h n o l o g y . S i n c e neurohormones a r e p r o t e i n s o r p e p t i d e s , t h e y a r e gene t r a n s l a t i o n p r o d u c t s and a r e amenable t o r e c o m b i n a n t DNA t e c h n o l o g y and g e n e t i c e n g i n e e r i n g . The most o b v i o u s appproach t o u s i n g recombinant DNA t e c h n o l o g y i s t o e n g i n e e r a neurohormone gene d i r e c t l y i n t o the genome o f the h o s t p l a n t f o r a pest insect. In t h i s manner, the h o s t p l a n t would t h e o r e t i c a l l y produce the hormone and p e s t s t h a t f e e d on the g e n e t i c a l l y t r a n s f o r m e d p l a n t s would r e c e i v e e x c e s s i v e doses o f the hormone r e s u l t i n g i n hyperstimulation of s e n s i t i v e processes. Other approaches f o r d e l i v e r i n g neurohormones by g e n e t i c e n g i n e e r i n g a r e t o t r a n s f o r m m i c r o o r g a n i s m s such as b a c t e r i a , p r o t o z o a o r y e a s t s w i t h neurohormone g e n e s . I n a l l o f the above c a s e s , the hormone i s a v a i l a b l e t o the i n s e c t by consuming the g e n e t i c a l l y - e n g i n e e r e d vector. However, d i g e s t i v e d e g r a d a t i o n o f the n a t u r a l hormone s t i l l remains as a problem s i n c e the hormone must pass the a l i m e n t a r y system o f the i n s e c t . F u r t h e r m o r e , s i n c e neurohormones a r e s y n t h e s i z e d as prohormones and a r e p o s t - t r a n s l a t i o n a l l y p r o c e s s e d i n t o a c t i v e hormones, i t remains t o be p r o v e n whether the p r o d u c t s o f neurohormone genes can be p r o c e s s e d a p p r o p r i a t e l y i n the c e l l s o f p l a n t s , b a c t e r i a , or protozoa. I t might be p o s s i b l e t o s y n t h e s i z e the s h o r t DNA sequence t h a t t r a n s c r i b e s s p e c i f i c a l l y f o r the amino a c i d sequence o f the a c t i v e hormone, and e n g i n e e r t h i s s h o r t "gene" i n t o a p l a n t o r c l o n i n g - e x p r e s s i o n v e c t o r so t h a t the DNA i s t r a n s c r i b e d and the hormone t r a n s l a t e d . Whether s h o r t segments o f DNA t h a t encode d i r e c t l y f o r the a c t i v e hormone can be e x p r e s s e d remains t o be d e m o n s t r a t e d . I n s e c t b a c u l o v i r u s e s a r e an a l t e r n a t i v e t o p l a n t s o r s i n g l e - c e l l e d organisms as c l o n i n g - e x p r e s s i o n v e c t o r s f o r neurohormone g e n e s . B a c u l o v i r u s e s a r e f r e q u e n t d i s e a s e a g e n t s o f l e p i d o p t e r a l a r v a e and were c o n s i d e r e d a t one time as h a v i n g a h i g h



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p o t e n t i a l as b i o c o n t r o l a g e n t s f o r p e s t i n s e c t s . R e c e n t l y , the b a c u l o v i r u s o f Autographa c a l i f o r n i c a has been g e n e t i c a l l y e n g i n e e r e d t o s e r v e as a c l o n i n g - e x p r e s s i o n v e c t o r f o r f o r e i g n genes (32)· T r a n s f o r m a t i o n o f the b a c u l o v i r u s e x p r e s s i o n v e c t o r (BEV) w i t h the human b e t a - i n t e r f e r o n gene f o l l o w e d by i n f e c t i o n o f i n s e c t c e l l c u l t u r e s r e s u l t e d i n the p r o d u c t i o n and s e c r e t i o n by i n f e c t e d c e l l s o f l a r g e amounts o f b i o l o g i c a l l y - a c t i v e i n t e r f e r o n . The a b i l i t y o f BEVs t o produce f o r e i g n gene p r o d u c t s JLn v i v o was c o n f i r m e d u s i n g the b a c u l o v i r u s o f B. m o r i s i l k w o r m s t r a n s f o r m e d w i t h the human a l p h a - i n t e r f e r o n gene. In t h i s c a s e , a l p h a - i n t e r f e r o n was found i n the hemolymph o f s i l k w o r m l a r v a e i n f e c t e d w i t h the recombinant BEV ( 3 3 ) . These f i n d i n g s demonstrate t h a t n u c l e a r p o l y h e d r o s i s v i r u s e s can c a r r y f o r e i g n genes i n t o i n f e c t e d l a r v a e , and the genes a r e e x p r e s s e d and the t r a n s l a t i o n products processed i n t o b i o l o g i c a l l y - a c t i v e molecules that are r e l e a s e d from i n f e c t e d c e l l s i n t o the h o s t ' s c i r c u l a t o r y system. Based on t h i s , i t i s t h e o r e t i c a l l y p o s s i b l e t h a t BEV g e n e t i c a l l y - e n g i n e e r e d w i t h neurohormone genes would c a r r y the genes i n t o h o s t i n s e c t s and e x p r e s s the hormonal p e p t i d e p r o d u c t s i n uncontrolled excess. The hormones would by-pass the d i g e s t i v e system, be r e l e a s e d d i r e c t l y i n t o the c i r c u l a t o r y system and would cause h y p e r s t i m u l a t i o n of s e n s i t i v e p h y s i o l o g i c a l processes, f o r example water e x c r e t i o n which would cause r a p i d d e h y d r a t i o n and d e a t h o f the i n f e c t e d l a r v a . R e c o m b i n a t i o n w i t h a p p r o p r i a t e f o r e i g n genes c o u l d make the v i r u s a more e f f e c t i v e b i o c o n t r o l a g e n t . BEV p a t h o l o g y i s slow and o f t e n r e q u i r e s n e a r l y a week t o k i l l i n f e c t e d i n s e c t s . A l s o , t h e r e i s a dose-dependency between the number o f i n g e s t e d v i r u s c a p s u l e s and l a r v a l s i z e . A n e u r o e n d o c r i n e p r o d u c t produced by a t r a n s f o r m e d v i r u s might improve the v i r u s as a b i o c o n t r o l a g e n t by i n h i b i t i n g f e e d i n g and k i l l i n g an i n f e c t e d l a r v a e a r l y , b e f o r e the o n s e t o f the v i r a l p a t h o l o g y , or by c o n v e r t i n g a s u b l e t h a l i n f e c t i o n i n t o a f a t a l i n f e c t i o n due t o p h y s i o l o g i c a l weakening and i n c r e a s e d susceptibility. A BEV t r a n s f o r m e d w i t h a neurohormone gene has the advantage t h a t i t i s an i n s e c t - s p e c i f i c b i o l o g i c a l agent combined w i t h a n a t u r a l , i n s e c t b i o r a t i o n a l f a c t o r . Such a c o m b i n a t i o n s h o u l d be o f m i n i m a l e n v i r o n m e n t a l c o n c e r n . I t i s a l s o f e a s i b l e t o t r a n s f o r m BEVs w i t h n a t u r a l or s y n t h e t i c genes f o r p r o t e i n a c e o u s a g e n t s t h a t would modulate neurohormone balance. For example, i t i s r e p o r t e d t h a t the a n t i s e n s e s t r a n d o f DNA c a r r i e s a g e n e t i c code t h a t i s complementary t o the sense s t r a n d (34)· Where the sense s t r a n d codes f o r h y d r o p h o b i c amino a c i d s , the a n t i s e n s e s t r a n d codes f o r h y d r o p h i l i c amino a c i d s , and v i c e v e r s a . N e u t r a l amino a c i d s p a i r g e n e r a l l y w i t h n e u t r a l amino a c i d s . The amino a c i d sequence was s y n t h e s i z e d f o r a p e p t i d e complementary t o ACTH as based on the a n t i s e n s e code f o r ACTH-mRNA (35). The r e s u l t i n g a n t i - A C T H mimicked a r e c e p t o r p r o t e i n and bound s e l e c t i v e l y t o ACTH. I f t h i s p a t t e r n o f p e p t i d e c o m p l e m e n t a r i t y i s v a l i d , then once i n s e c t neurohormone genes are i s o l a t e d and t h e i r base sequences d e t e r m i n e d , i t s h o u l d be p o s s i b l e t o s y n t h e s i z e anti-hormone genes, c l o n e them i n t o a BEV and produce an antihormone t h a t w i l l t i e - u p the n a t u r a l hormone a t a c r i t i c a l time i n the l i f e o f the i n s e c t ( e . g . c u t i c l e t a n n i n g , e c l o s i o n ) . T h i s t e c h n o l o g y would be most u s e f u l w i t h the l a r g e m o l e c u l a r w e i g h t neurohormones such as PTTH, e c l o s i o n hormone o r b u r s i c o n .


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S e v e r a l o t h e r a l t e r n a t i v e s may be p o s s i b l e u s i n g g e n e t i c engineering. For example, i t may be p o s s i b l e t o d e s i g n a r t i f i c i a l p r o t e i n s and t h e i r a s s o c i a t e d genes t h a t w i l l a c t as hormone a n t a g o n i s t s or mimics a t hormone a c c e p t o r s i t e s ( t a r g e t c e l l , degradative proteases). F i n a l l y , i t may be p o s s i b l e t o produce m o n o c l o n a l a n t i b o d i e s t o a g i v e n hormone, i s o l a t e the gene f o r the a n t i b o d y , i n s e r t the a n t i b o d y gene i n t o the BEV and e x p r e s s an a n t i b o d y t h a t would b i n d and f u n c t i o n a l l y remove s p e c i f i c hormones from B E V - i n f e c t e d i n s e c t s . Altered Neurosecretion. B i o g e n i c amines a f f e c t s p e c i f i c hormone s e c r e t i o n i n the n e u r o e n d o c r i n e system o f v e r t e b r a t e s . The b i o g e n i c amines may e i t h e r i n h i b i t or s t i m u l a t e n e u r o s e c r e t i o n d i r e c t l y (_36), o r t h e y may r e g u l a t e n e u r o s e c r e t i o n i n d i r e c t l y t h r o u g h c o n t r o l s o v e r h o r m o n e - s p e c i f i c r e l e a s i n g f a c t o r s (^8)· B i o g e n i c amines a l s o a f f e c t n e u r o s e c r e t i o n i n i n s e c t s . For example, r e s e r p i n e d e p l e t e s b i o g e n i c amines from the nervous system, and t r e a t m e n t o f P. a m e r i c a n a w i t h r e s e r p i n e s t i m u l a t e s the s e c r e t i o n o f h y p e r g l y c e m i c hormones (37) · T h i s r e s u l t s u g g e s t s t h a t the b i o g e n i c amines i n h i b i t h y p e r g l y c e m i c hormone s e c r e t i o n . L o c u s t s p o i s o n e d w i t h i n s e c t i c i d e s t h a t a c t on the c h o l i n e r g i c nervous system show enhanced ΑΚΗ s e c r e t i o n (_38). However, ΑΚΗ s e c r e t i o n i s s u p p r e s s e d i f the t e s t i n s e c t s a r e p r e t r e a t e d w i t h a g e n t s t h a t i n h i b i t the a m i n e r g i c nervous system b e f o r e t h e y a r e exposed t o the i n s e c t i c i d e s (39,40)· These f i n d i n g s s u g g e s t t h a t the c h o l i n e r g i c nervous system i n f l u e n c e s the a m i n e r g i c nervous system which, i n t u r n , r e g u l a t e s s p e c i f i c neurosecretion. Of the i n s e c t b i o g e n i c amines, octopamine appears t o e x e r t the most e f f e c t on n e u r o s e c r e t i o n (41_,42). Octopamine has l i t t l e s i g n i f i c a n c e i n the v e r t e b r a t e nervous system where i t i s e i t h e r degraded r a p i d l y or may s e r v e as a c o t r a n s m i t t e r w i t h norepinephrine. Exposure o f c o r p o r a c a r d i a c a t o octopamine i n v i t r o promotes ΑΚΗ s e c r e t i o n ( 4J_), and c h l o r d i m e f orm, an i n s e c t i c i d a l a g e n t and octopamine mimic, a l s o s t i m u l a t e s ΑΚΗ s e c r e t i o n from i s o l a t e d c o r p o r a c a r d i a c a (43). This l a t t e r f i n d i n g i l l u s t r a t e s t h a t c h e m i c a l s can be a p p l i e d t o i n s e c t s w i t h the e x p e c t a t i o n t h a t they w i l l a f f e c t or mimic the a m i n e r g i c nervous system o f i n s e c t s t o r e g u l a t e s p e c i f i c n e u r o s e c r e t i o n . More f u n d a m e n t a l i n f o r m a t i o n about the r e g u l a t i o n o f n e u r o s e c r e t o r y p r o c e s s e s i n i n s e c t s may s u g g e s t ways t o m a n i p u l a t e n e u r o s e c r e t i o n f o r pragmatic purposes. I n h i b i t i o n o f Enzymes f o r Hormone S y n t h e s i s or D e g r a d a t i o n . The q u e s t i o n of i n h i b i t i n g enzymes t h a t are i n s t r u m e n t a l i n e i t h e r the s y n t h e s i s or d e g r a d a t i o n o f i n s e c t neurohormones i s d i f f i c u l t t o a d d r e s s a t t h i s t i m e . L i t t l e i s known about the s y n t h e t i c enzymes o f v e r t e b r a t e neurohormones and n o t h i n g i s known about the i n s e c t enzymes. T h i s a r e a must a w a i t more r e s e a r c h i n the f i e l d o f b o t h animal groups. The y e a s t KEX2 p r o t e a s e has the p r o p e r t i e s o f a p r o h o r m o n e - c o n v e r t i n g enzyme (44) and may p r o v i d e a model f o r s t u d y i n g prohormone p r o c e s s i n g . Captopril, i l l u s t r a t e s that i t i s p o s s i b l e t o m a n i p u l a t e p e p t i d e hormone t i t e r s by i n h i b i t i n g a p p r o p r i a t e s y n t h e t i c o r a c t i v a t i n g enzymes. C a p t o p r i l i s a s y n t h e t i c o r a l a n t i h y p e r t e n s i v e t h a t was d e s i g n e d s p e c i f i c a l l y t o i n h i b i t the a n g i o t e n s i n - c o n v e r t i n g enzyme t h a t c o n v e r t s i n a c t i v e


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a n g i o t e n s i n I t o a c t i v e a n g i o t e n s i n I I {45)· Alternatively, l i t t l e i s a l s o knovm about the d e g r a d a t i v e enzymes f o r i n s e c t neurohormones. Aminopeptidase was i d e n t i f i e d as b e i n g i m p o r t a n t f o r i n v i v o i n a c t i v a t i o n of p r o c t o l i n i n cockroaches (46,47). The a b i l i t y t o a l t e r hormone t i t e r s by a l t e r i n g the s y n t h e s i s and d e g r a d a t i o n o f i n s e c t neurohormones a w a i t s more b a s i c r e s e a r c h , s p e c i f i c a l l y : c h e m i c a l d e f i n i t i o n o f i n s e c t neurohormones; i s o l a t i o n o f the genes and prohormone forms i n v o l v e d i n the s y n t h e s i s o f the d e f i n e d hormones; and p r e p a r a t i o n o f l a b e l e d hormones f o r use i n degradation s t u d i e s . Neurohormone A n a l o g s . F i n a l l y , a n a l o g s o f the neurohormones p r o b a b l y have the g r e a t e s t p o t e n t i a l f o r m a n i p u l a t i n g neuroendocrine-regulated processes. There a r e s e v e r a l r e a s o n s why a n a l o g s a r e the most f e a s i b l e a p p r o a c h t o a n e u r o e n d o c r i n e - b a s e d strategy for pest insect c o n t r o l . F i r s t , there i s a long h i s t o r y of e x p e r i e n c e and s u c c e s s c o n c e r n e d w i t h the use o f c h e m i c a l s t o control insect pests. Second, a n a l o g s have a n e a r l y i n f i n i t e p o t e n t i a l f o r m o d i f i c a t i o n s t o improve s t a b i l i t y , p e n e t r a b i l i t y , t o x i c i t y and s p e c i f i c i t y . We know t h a t t h e r e a r e numerous i n s e c t neurohormones and t h a t s e v e r a l forms may e x i s t f o r the same hormone, b o t h between and w i t h i n i n s e c t s p e c i e s . T h e r e f o r e , neurohormones p r e s e n t the s y n t h e t i c p e p t i d e c h e m i s t w i t h a m u l t i t u d e o f p o s s i b i l i t i e s f o r use i n d e s i g n i n g i n s e c t i c i d a l a g e n t s . I t i s argued t h a t the commercial manufacture o f p e p t i d e s i s t o o expensive f o r a g r i c u l t u r a l use. However, I do n o t propose t h a t the f i n a l a n a l o g w i l l be a p e p t i d e . R a t h e r , the p r a c t i c a l agent w i l l p r o b a b l y be a n o n p e p t i d i c d e r i v a t i v e o f a neurohormone t h a t i s l o c k e d i n t o a s t r u c t u r e t h a t mimics the a c t i v e c o n f o r m a t i o n o f the hormone. H i s t o r i c a l l y , i f an agent i s s u f f i c i e n t l y e f f e c t i v e and an adequate market e x i s t s , then i n d u s t r y w i l l s o l v e the problems o f e c o n o m i c a l p r o d u c t i o n and a p p l i c a t i o n . A u s e f u l a n a l o g f o r a p e p t i d e may no l o n g e r be p e p t i d i c . There i s not s u f f i c i e n t space t o d e s c r i b e the d e t a i l s o f how an a n a l o g can be d e s i g n e d f o r a g i v e n neurohormone. Furthermore, each p e p t i d e p o s s e s s e s unique p r o p e r t i e s t h a t r e q u i r e i n d i v i d u a l approaches f o r a n a l o g d e s i g n , and more d e t a i l s r e l a t e d t o a n a l o g d e s i g n were p r e s e n t e d i n a s e p a r a t e paper (J_£) · What i s modeled i n d e s i g n i n g an a n a l o g i s the b i o l o g i c a l l y - a c t i v e c o n f o r m a t i o n assumed by the p e p t i d e as i t i n t e r a c t s w i t h i t s t a r g e t - c e l l r e c e p t o r . A p e p t i d e hormone c a r r i e s two t y p e s o f i n f o r m a t i o n : an a d d r e s s and a message. The a d d r e s s p o r t i o n o f the p e p t i d e i s t h a t r e g i o n t h a t f i t s i n t o the c o r r e c t r e c e p t o r . The message i s e i t h e r p a r t o f the a d d r e s s or i t can be a s e p a r a t e r e g i o n , and i t a c t i v a t e s the r e c e p t o r t o i n i t i a t e the c e l l u l a r r e s p o n s e . A n t a g o n i s t i c a n a l o g s c o n t a i n o n l y an a d d r e s s , and a n t a g o n i s t s f i t i n t o the r e c e p t o r w i t h o u t c o n v e y i n g a message. A l t e r n a t i v e l y , a g o n i s t i c a n a l o g s c o n t a i n both a f u n c t i o n a l a d d r e s s and message and produce a c e l l u l a r r e s p o n s e . The d e s i r e o f the a n a l o g a p p r o a c h i s t o d e v e l o p m o l e c u l e s t h a t a r e s t a b l e and l o c k e d i n t o the a c t i v e c o n f o r m a t i o n so t h a t the a n a l o g e i t h e r b i n d s the r e c e p t o r and p r e v e n t s the n a t u r a l hormone from e x e r t i n g i t s a c t i o n (an a n t a g o n i s t ) , o r the a n a l o g b i n d s the r e c e p t o r and causes h y p e r s t i m u l a t i o n o f the p h y s i o l o g i c a l r e s p o n s e (a s u p e r a g o n i s t ) . The a n a l o g i s e i t h e r s t a b i l i z e d t o r e s i s t enzymic d e g r a d a t i o n o r the enzyme does n o t a c t on the a n a l o g because o f i t s n o n p e p t i d i c n a t u r e .



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These p r o p e r t i e s p e r m i t the a n a l o g t o m a i n t a i n a c o n t i n u o u s l y h i g h t i t e r and t o s u s t a i n i t s e f f e c t . Several p h y s i o l o g i c a l processes are s u s c e p t i b l e t o superagonists. F o r example, a s u p e r a g o n i s t t o t h e d i u r e t i c hormone c o u l d cause s e v e r e d e h y d r a t i o n , i m m o b i l i t y and d e a t h i n l a r v a l i n s e c t s , e s p e c i a l l y those i n a d r y e n v i r o n m e n t such as s t o r e d g r a i n . H y p e r s t i m u l a t i o n o f JH p r o d u c t i o n by an a l l a t o t r o p i n a g o n i s t might r e s u l t i n a b n o r m a l l y h i g h l e v e l s o f J H a t t h e time f o r e p i d e r m a l c e l l recommitment, hence metamorphosis would n o t o c c u r . Conversely, an a l l a t o s t a t i n a g o n i s t would i n h i b i t JH p r o d u c t i o n and promote premature metamorphosis. A n t a g o n i s t i c a n a l o g s t o a l l a t o t r o p i n and a l l a t o s t a t i n c o u l d be used i n a r e c i p r o c a l manner t o a g o n i s t s f o r r e g u l a t i n g JH p r o d u c t i o n . An a n t a g o n i s t t o a l l a t o t r o p i n would i n h i b i t JH s y n t h e s i s and promote premature metamorphosis; whereas, an a l l a t o s t a t i n i n h i b i t o r would promote J H s y n t h e s i s and p r e v e n t metamorphosis. F i n a l l y , a n a l o g s might be u s e f u l f o r d i s r u p t i n g t h e a c t i o n o f myokinins. A n t a g o n i s t i c analogs t o myokinins t h a t a f f e c t gut c o n t r a c t i l i t y might s u p p r e s s f e e d i n g a c t i v i t y , and a n a l o g s t o n e u r o p e p t i d e s t h a t a f f e c t s k e l e t a l muscles might r e s u l t i n p a r a l y s i s and i m m o b i l i t y . I f t h e a n a l o g s were s t a b l e t o enzymic d e g r a d a t i o n , t h e y might remain a c t i v e f o r an extended time and cause s u s t a i n e d p a r a l y s i s and d e a t h . Summary I t i s my i n t e n t t h a t t h i s d i s c u s s i o n s h o u l d s t i m u l a t e c o n s i d e r a t i o n of the p r a c t i c a l p o t e n t i a l f o r i n s e c t neuroendocrine r e s e a r c h . I t i s u n l i k e l y t h a t we w i l l d e v e l o p p r a c t i c a l neurohormone-based s t r a t e g i e s f o r i n s e c t p e s t management i n t h e near f u t u r e . C o n s e r v a t i v e l y , I would p r o j e c t t h a t any pragmatic use o f c u r r e n t n e u r o e n d o c r i n e r e s e a r c h would be f i v e y e a r s away a t t h e e a r l i e s t and more on the o r d e r o f t e n o r more y e a r s . A t p r e s e n t , we need more fundamental i n f o r m a t i o n on t h e c h e m i c a l n a t u r e o f t h e i n s e c t neurohormones, t h e i r e n d o c r i n o l o g y and t h e i r p h y s i o l o g i c a l s i g n i f i c a n c e w i t h i n the i n s e c t s . I t i s e s s e n t i a l t h a t we s t r u c t u r a l l y d e f i n e more neurohormones and d e v e l o p s p e c i f i c a s s a y s f o r t h e s e hormones i n o r d e r t o s t u d y t h e i r e n d o c r i n o l o g y . Finally, we need t o d e t e r m i n e what s i g n i f i c a n c e t h e d i s r u p t i o n o f a p h y s i o l o g i c a l p r o c e s s would have on s u r v i v a l . W i t h o u t t h i s base o f fundamental i n f o r m a t i o n , i t w i l l n o t be p o s s i b l e t o d e v e l o p the types o f pragmatic s t r a t e g i e s proposed here. Acknowledgments P e r s o n a l r e s e a r c h r e l a t e d t o t h i s p u b l i c a t i o n was s u p p o r t e d b y NSF g r a n t no. DCB 8511058 and by t h e Robert J . K l e b e r g , J r . and Helen C. Kleberg Foundation.

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Kopec, S. Bull. Int. Acad. Cracovie 1917, B, 57-60. Scharrer, B. J. Comp. Neurol. 1941, 74, 93-108. Raabe, M. In Insect Neurohormones; Plenum: NY, 1982. Starratt, A.N.; Brown, B.E. Life Sci. 1975,17,1253-1256.


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